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Sample records for acidic protein levels

  1. Maternal folic acid supplementation to dams on marginal protein level alters brain fatty acid levels of their adult offspring.

    PubMed

    Rao, Shobha; Joshi, Sadhana; Kale, Anvita; Hegde, Mahabaleshwar; Mahadik, Sahebarao

    2006-05-01

    Studies on fetal programming of adult diseases have highlighted the importance of maternal nutrition during pregnancy. Folic acid and long-chain essential polyunsaturated fatty acids (LC-PUFAs) have independent effects on fetal growth. However, folic acid effects may also involve alteration of LC-PUFA metabolism. Because marginal deficiency of LC-PUFAs during critical periods of brain growth and development is associated with risks for adult diseases, it is highly relevant to investigate how maternal supplementation of such nutrients can alter brain fatty acid levels. We examined the impact of folic acid supplementation, conventionally used in maternal intervention, on brain essential fatty acid levels and plasma corticosterone concentrations in adult offspring at 11 months of age. Pregnant female rats from 4 groups (6 in each) were fed with casein diets either with 18 g protein/100 g diet (control diet) or treatment diets that were marginal in protein (MP), such as 12 g protein/100 g diet supplemented with 8 mg folic acid (FAS/MP), 12 g protein/100 g diet without folic acid (FAD/MP), or 12 g protein/100 g diet (MP) with 2 mg folic acid. Pups were weaned to a standard laboratory diet with 18 g protein/100 g diet. All male adult offspring in the FAS/MP group showed lower docosahexaenoic acid (P<.05) as compared with control adult offspring (6.04+/-2.28 vs 10.33+/-0.86 g/100 g fatty acids) and higher n-6/n-3 ratio (P<.05). Docosahexaenoic acid levels in FAS/MP adult offspring were also lower (P<.05) when compared with the MP group. Plasma corticosterone concentrations were higher (P<.05) in male adult offspring from the FAS/MP group compared with control as well as the MP adult offspring. Results suggest that maternal folic acid supplementation at MP intake decreased brain docosahexaenoic acid levels probably involving corticosterone increase. PMID:16631439

  2. Maternal folic acid supplementation to dams on marginal protein level alters brain fatty acid levels of their adult offspring.

    PubMed

    Rao, Shobha; Joshi, Sadhana; Kale, Anvita; Hegde, Mahabaleshwar; Mahadik, Sahebarao

    2006-05-01

    Studies on fetal programming of adult diseases have highlighted the importance of maternal nutrition during pregnancy. Folic acid and long-chain essential polyunsaturated fatty acids (LC-PUFAs) have independent effects on fetal growth. However, folic acid effects may also involve alteration of LC-PUFA metabolism. Because marginal deficiency of LC-PUFAs during critical periods of brain growth and development is associated with risks for adult diseases, it is highly relevant to investigate how maternal supplementation of such nutrients can alter brain fatty acid levels. We examined the impact of folic acid supplementation, conventionally used in maternal intervention, on brain essential fatty acid levels and plasma corticosterone concentrations in adult offspring at 11 months of age. Pregnant female rats from 4 groups (6 in each) were fed with casein diets either with 18 g protein/100 g diet (control diet) or treatment diets that were marginal in protein (MP), such as 12 g protein/100 g diet supplemented with 8 mg folic acid (FAS/MP), 12 g protein/100 g diet without folic acid (FAD/MP), or 12 g protein/100 g diet (MP) with 2 mg folic acid. Pups were weaned to a standard laboratory diet with 18 g protein/100 g diet. All male adult offspring in the FAS/MP group showed lower docosahexaenoic acid (P<.05) as compared with control adult offspring (6.04+/-2.28 vs 10.33+/-0.86 g/100 g fatty acids) and higher n-6/n-3 ratio (P<.05). Docosahexaenoic acid levels in FAS/MP adult offspring were also lower (P<.05) when compared with the MP group. Plasma corticosterone concentrations were higher (P<.05) in male adult offspring from the FAS/MP group compared with control as well as the MP adult offspring. Results suggest that maternal folic acid supplementation at MP intake decreased brain docosahexaenoic acid levels probably involving corticosterone increase.

  3. Ileal apical sodium-dependent bile acid transporter protein levels are down-regulated through ubiquitin-dependent protein degradation induced by bile acids.

    PubMed

    Miyata, Masaaki; Yamakawa, Hiroki; Hayashi, Kenjiro; Kuribayashi, Hideaki; Yamazoe, Yasushi; Yoshinari, Kouichi

    2013-08-15

    The ileal apical sodium-dependent bile acid transporter (ASBT or SLC10A2) has a crucial role in intestinal bile acid absorption. We previously reported that enterobacteria-mediated bile acid conversion was involved in the alteration of ileal ASBT expression levels. In the present study, to investigate the hypothesis that ileal ASBT protein levels are post-translationally regulated by enterobacteria-associated bile acids, alteration of ileal ASBT protein levels was analysed in mice 12 h and 24 h after anti-bacterial drug ampicillin (ABPC) treatment (100 mg/kg, single shot) that altered bile acid composition in the intestinal lumen. In ABPC-treated mice, enterobacteria-biotransformed bile acid, taurodeoxycholic acid (TDCA) and cholic acid (CA) levels were decreased, whereas taurocholic acid (TCA) and tauro-β-muricholic acid levels were increased in the intestinal lumen. Ileal ASBT protein levels in brush-border membrane vesicles (BBMVs), but not ileal Asbt mRNA levels, were significantly increased in the ABPC-treated mice, and the extent of ubiquitination of the ileal ASBT protein was reduced in the ABPC-treated mice. Treatment of ABPC-pretreated mice with CA or TDCA, but not TCA, significantly decreased ileal ASBT protein levels and increased the extent of ubiquitination of ileal ASBT protein. Treatment of mice with the lysosome inhibitor, chloroquine, or the proteasome inhibitor, MG132, increased ileal ASBT protein levels in BBMVs. CA-mediated reduction of ASBT protein levels in the ABPC-pretreated mice was attenuated by co-treatment with chloroquine or MG132. These results suggest that ileal ASBT protein is degraded by a ubiquitin-dependent pathway in response to enterobacteria-associated bile acids. PMID:23872411

  4. Elevated glial fibrillary acidic protein levels in the cerebrospinal fluid of patients with narcolepsy.

    PubMed

    Feneberg, Emily; Steinacker, Petra; Lehnert, Stefan; Böhm, Bernhard; Mayer, Geert; Otto, Markus

    2013-07-01

    Glial fibrillary acidic protein (GFAP) is an established indicator of astrogliosis. Therefore, variable cerebrospinal fluid (CSF) concentrations of this protein might reflect disease-specific pathologic profiles. In patients with narcolepsy, a loss of hypocretin-1 (hcrt-1) neurons in the brain and low concentrations of hcrt-1 in CSF have been reported. We performed a commercially available enzyme-linked immunosorbent assay to investigate if GFAP also is altered in the CSF of these patients. Here we detected significantly higher CSF levels of GFAP in patients with low hcrt-1 levels, of which the majority had a diagnosis of narcolepsy and cataplexy (NC); however, this finding was not observed in patients with hcrt-1 levels that were within reference range. In conclusion, GFAP may be useful as an additional disease biomarker in patients with narcolepsy, and this hypothesis should be investigated in larger studies.

  5. Association of androgen with gender difference in serum adipocyte fatty acid binding protein levels

    PubMed Central

    Hu, Xiang; Ma, Xiaojing; Pan, Xiaoping; Luo, Yuqi; Xu, Yiting; Xiong, Qin; Bao, Yuqian; Jia, Weiping

    2016-01-01

    Clinical investigations have indicated women have higher levels of adipocyte fatty acid binding protein (A-FABP) than men. The present study aimed to identify factors related to gender difference in serum A-FABP levels. A total of 507 participants (194 men, 132 premenopausal women, and 181 postmenopausal women) were enrolled in the present study. Serum A-FABP levels increased in the order from men to premenopausal women to postmenopausal women in both body mass index categories (<25.0 and ≥25.0 kg/m2; all P < 0.05). Multiple stepwise regression analyses showed that after adjustment for factors related to serum A-FABP levels, the trunk fat mass was an independent and positive factor of serum A-FABP levels. For men, total testosterone was associated independently and inversely with serum A-FABP levels. For pre- and postmenopausal women, bioavailable testosterone and total testosterone were independent and positive factors associated with serum A-FABP levels, respectively. The present study demonstrated that the androgen was correlated with the serum A-FABP levels negatively in men, but positively in women. With these effects on the fat content, especially trunk fat, androgen might contribute to the gender difference in serum A-FABP levels. PMID:27270834

  6. Uncoupling protein 3 expression levels influence insulin sensitivity, fatty acid oxidation, and related signaling pathways.

    PubMed

    Senese, Rosalba; Valli, Vivien; Moreno, Maria; Lombardi, Assunta; Busiello, Rosa Anna; Cioffi, Federica; Silvestri, Elena; Goglia, Fernando; Lanni, Antonia; de Lange, Pieter

    2011-01-01

    Controversy exists on whether uncoupling protein 3 (UCP3) positively or negatively influences insulin sensitivity in vivo, and the underlying signaling pathways have been scarcely studied. We studied how a progressive reduction in UCP3 expression (using UCP3 +/+, UCP3 +/-, and UCP3 -/- mice) modulates insulin sensitivity and related metabolic parameters. In order to further validate our observations, we also studied animals in which insulin resistance was induced by administration of a high-fat diet (HFD). In UCP3 +/- and UCP3 -/- mice, gastrocnemius muscle Akt/protein kinase B (Akt/PKB) (serine 473) and AMP-activated protein kinase (AMPK) (threonine 171) phosphorylation, and glucose transporter 4 (GLUT4) membrane levels were reduced compared to UCP3 +/+ mice. The HOMA-IR index (insulin resistance parameter) was increased both in the UCP3 +/- and UCP3 -/- mice. In these mice, insulin administration normalized Akt/PKB phosphorylation between genotypes while AMPK phosphorylation was further reduced, and sarcolemmal GLUT4 levels were induced but did not reach control levels. Furthermore, non-insulin-stimulated muscle fatty acid oxidation and the expression of several involved genes both in muscle and in liver were reduced. HFD administration induced insulin resistance in UCP3 +/+ mice and the aforementioned parameters resulted similar to those of chow-fed UCP3 +/- and UCP3 -/- mice. In conclusion, high-fat-diet-induced insulin resistance in wild-type mice mimics that of chow-fed UCP3 +/- and UCP3 -/- mice showing that progressive reduction of UCP3 levels results in insulin resistance. This is accompanied by decreased fatty acid oxidation and a less intense Akt/PKB and AMPK signaling.

  7. Amino acids and proteins.

    PubMed

    van Goudoever, Johannes B; Vlaardingerbroek, Hester; van den Akker, Chris H; de Groof, Femke; van der Schoor, Sophie R D

    2014-01-01

    Amino acids and protein are key factors for growth. The neonatal period requires the highest intake in life to meet the demands. Those demands include amino acids for growth, but proteins and amino acids also function as signalling molecules and function as neurotransmitters. Often the nutritional requirements are not met, resulting in a postnatal growth restriction. However, current knowledge on adequate levels of both amino acid as well as protein intake can avoid under nutrition in the direct postnatal phase, avoid the need for subsequent catch-up growth and improve later outcome.

  8. Effects of copper on chlorophyll, proline, protein and abscisic acid level of sunflower (Helianthus annuus L.) seedlings.

    PubMed

    Zengin, Fikriye Kirbag; Kirbag, Sevda

    2007-07-01

    The effect of copperchloride (CuCl2) on the level of chlorophyll (a+b), proline, protein and abscisic acid in sunflower (Helianthus annuus L.) seedlings were investigated Control and copper treated (0.4, 0.5 and 0.6 mM) seedlings were grown for ten days in Hoagland solution. Abscisic acid content was determined in root, shoot and leaf tissues of seedlings by HPLC. Copper stress caused significant increase of the abscisic acid contents in roots, shoots and leaves of seedlings. The increase was dependent on the copper salt concentration. Enhanced accumulation of proline in the leaves of seedlings exposed to copper was determined, as well as a decrease of chlorophyll (a+b) and total protein (p < 0.05 or p < 0.01). It was observed that the level of chlorophyll (a+b) and total protein (p < 0.05 or p < 0.01) remarkably decreased as copper concentration increased to 0.6 mM, although the levels of proline and abscisic acid in the leaves of plants were increased--a dose-depended behavior The same trends were also observed with the level of abscisic acid of stems and roots. Copper has dose- depended effects on chlorophyll, proline, protein and abscisic acid level of sunflower (Helianthus annuus L.) seedlings. Thus, we assumed that copper levels increase above some critical points seedling growth get negative effects. This assumption is in line with previous findings.

  9. ABI4 mediates antagonistic effects of abscisic acid and gibberellins at transcript and protein levels.

    PubMed

    Shu, Kai; Chen, Qian; Wu, Yaorong; Liu, Ruijun; Zhang, Huawei; Wang, Pengfei; Li, Yanli; Wang, Shengfu; Tang, Sanyuan; Liu, Chunyan; Yang, Wenyu; Cao, Xiaofeng; Serino, Giovanna; Xie, Qi

    2016-02-01

    Abscisic acid (ABA) and gibberellins (GAs) are plant hormones which antagonistically mediate numerous physiological processes, and their optimal balance is essential for normal plant development. However, the molecular mechanism underlying ABA and GA antagonism still needs to be determined. Here, we report that ABA-INSENSITIVE 4 (ABI4) is a central factor in GA/ABA homeostasis and antagonism in post-germination stages. ABI4 overexpression in Arabidopsis (OE-ABI4) leads to developmental defects including a decrease in plant height and poor seed production. The transcription of a key ABA biosynthetic gene, NCED6, and of a key GA catabolic gene, GA2ox7, is significantly enhanced by ABI4 overexpression. ABI4 activates NCED6 and GA2ox7 transcription by directly binding to the promoters, and genetic analysis revealed that mutation in these two genes partially rescues the dwarf phenotype of ABI4 overexpressing plants. Consistently, ABI4 overexpressing seedlings have a lower GA/ABA ratio than the wild type. We further show that ABA induces GA2ox7 transcription while GA represses NCED6 expression in an ABI4-dependent manner; and that ABA stabilizes the ABI4 protein whereas GA promotes its degradation. Taken together, these results suggest that ABA and GA antagonize each other by oppositely acting on ABI4 transcript and protein levels.

  10. Ursolic acid regulates aging process through enhancing of metabolic sensor proteins level.

    PubMed

    Bahrami, Soroush Alaghehband; Bakhtiari, Nuredin

    2016-08-01

    We previously reported that Ursolic Acid (UA) ameliorates skeletal muscle performance through satellite cells proliferation and cellular energy status. In studying the potential role of the hypothalamus in aging, we developed a strategy to pursue UA effects on the hypothalamus anti-aging proteins such as; SIRT1, SIRT6, PGC-1β and α-Klotho. In this study, we used a model of aging animals (C57BL/6). UA dissolved in Corn oil (20mg/ml) and then administrated (200mg/Kg i.p injection) to mice, twice daily for 7days. After treatment times, the mice perfused and the hypothalamus isolated for preparing of tissue to Immunofluorescence microscopy. The data illustrated that UA significantly increased SIRT1 (∼3.5±0.3 folds) and SIRT-6 (∼1.5±0.2 folds) proteins overexpression (P<0.001). In addition, our results showed that UA enhanced α-Klotho (∼3.3±0.3) and PGC-1β (∼2.6±0.2 folds) proteins levels (P<0. 01). In this study, data were analyzed using SPSS 16 (ANOVA test). To the best of our knowledge, it seems that UA through enhancing of anti-aging biomarkers (SIRT1 and SIRT6) and PGC-1β in hypothalamus regulates aging-process and attenuates mitochondrial-related diseases. In regard to the key role of α-Klotho in aging, our data indicate that UA may be on the horizon to forestall diseases of aging. PMID:27470332

  11. Ursolic acid regulates aging process through enhancing of metabolic sensor proteins level.

    PubMed

    Bahrami, Soroush Alaghehband; Bakhtiari, Nuredin

    2016-08-01

    We previously reported that Ursolic Acid (UA) ameliorates skeletal muscle performance through satellite cells proliferation and cellular energy status. In studying the potential role of the hypothalamus in aging, we developed a strategy to pursue UA effects on the hypothalamus anti-aging proteins such as; SIRT1, SIRT6, PGC-1β and α-Klotho. In this study, we used a model of aging animals (C57BL/6). UA dissolved in Corn oil (20mg/ml) and then administrated (200mg/Kg i.p injection) to mice, twice daily for 7days. After treatment times, the mice perfused and the hypothalamus isolated for preparing of tissue to Immunofluorescence microscopy. The data illustrated that UA significantly increased SIRT1 (∼3.5±0.3 folds) and SIRT-6 (∼1.5±0.2 folds) proteins overexpression (P<0.001). In addition, our results showed that UA enhanced α-Klotho (∼3.3±0.3) and PGC-1β (∼2.6±0.2 folds) proteins levels (P<0. 01). In this study, data were analyzed using SPSS 16 (ANOVA test). To the best of our knowledge, it seems that UA through enhancing of anti-aging biomarkers (SIRT1 and SIRT6) and PGC-1β in hypothalamus regulates aging-process and attenuates mitochondrial-related diseases. In regard to the key role of α-Klotho in aging, our data indicate that UA may be on the horizon to forestall diseases of aging.

  12. DNA curtains: novel tools for imaging protein-nucleic acid interactions at the single-molecule level.

    PubMed

    Collins, Bridget E; Ye, Ling F; Duzdevich, Daniel; Greene, Eric C

    2014-01-01

    Interactions between proteins and nucleic acids are at the molecular foundations of most key biological processes, including DNA replication, genome maintenance, the regulation of gene expression, and chromosome segregation. A complete understanding of these types of biological processes requires tackling questions with a range of different techniques, such as genetics, cell biology, molecular biology, biochemistry, and structural biology. Here, we describe a novel experimental approach called "DNA curtains" that can be used to complement and extend these more traditional techniques by providing real-time information about protein-nucleic acid interactions at the level of single molecules. We describe general features of the DNA curtain technology and its application to the study of protein-nucleic acid interactions in vitro. We also discuss some future developments that will help address crucial challenges to the field of single-molecule biology.

  13. Proteomic Investigation of Protein Profile Changes and Amino Acid Residue Level Modification in Cooked Lamb Meat: The Effect of Boiling.

    PubMed

    Yu, Tzer-Yang; Morton, James D; Clerens, Stefan; Dyer, Jolon M

    2015-10-21

    Hydrothermal treatment (heating in water) is a common method of general food processing and preparation. For red-meat-based foods, boiling is common; however, how the molecular level effects of this treatment correlate to the overall food properties is not yet well-understood. The effects of differing boiling times on lamb meat and the resultant cooking water were here examined through proteomic evaluation. The longer boiling time was found to result in increased protein aggregation involving particularly proteins such as glyceraldehyde-3-phosphate dehydrogenase, as well as truncation in proteins such as in α-actinin-2. Heat-induced protein backbone cleavage was observed adjacent to aspartic acid and asparagine residues. Side-chain modifications of amino acid residues resulting from the heating, including oxidation of phenylalanine and formation of carboxyethyllysine, were characterized in the cooked samples. Actin and myoglobin bands from the cooked meat per se remained visible on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, even after significant cooking time. These proteins were also found to be the major source of observed heat-induced modifications. This study provides new insights into molecular-level modifications occurring in lamb meat proteins during boiling and a protein chemistry basis for better understanding the effect of this common treatment on the nutritional and functional properties of red-meat-based foods.

  14. Endogenous salicylic acid levels correlate with accumulation of pathogenesis-related proteins and virus resistance in tobacco

    SciTech Connect

    Yalpani, N.; Shulaev, V.; Raskin, I. )

    1993-07-01

    Salicylic acid (SA) is hypothesized to be an endogenous regulator of local and systemic disease resistance and an inducer of pathogenesis-related (PR) proteins among plants. High levels of PR proteins have been observed in an uninoculated amphidiploid hybrid of Nicotiana glutinosa [times] N. debneyi, which is highly resistant to tobacco mosaic virus (TMV). Fluoresence, UV, and mass spectral analysis established that the levels of SA in healthy N. glutinosa [times] N. debneyi leaves were 30 times greater than in N. tabacum [open quotes]Xanthi-nc[close quotes] tobacco, which does not constitutively express PR proteins and is less resistant to TMV. Upon TMV-inoculation SA levels increased at least 70-fold leaves of Xanthi-nc but role only slightly in the hybrid. Phloem exudates of N. glutinosa [times] N. debneyi contained at least 500 times more SA than those of Xanthi-nc. SA treatment caused the appearance of PR-1 protein in Xanthi-nc but did not affect constitutively high levels of PR-1 protein in N. glutinosa [times] N. debneyi. In contrast to Xanthi-nc tobacco, TMV-inoculated N. glutinosa [times] N. debneyi kept at 32 C accumulated more than 0.5 [mu]g SA/g fresh weight, maintained high levels of PR proteins, and developed a hypersensitive response to TMV. PR proteins have previously been shown to accumulate in the lower leaves of healthy, flowering Xanthi-nc tobacco, which exhibited increased resistance to TMV. These developmentally induced increases in resistance and PR-1 proteins positively correlated with tissue levels of SA. These results affirm the regulatory role of SA in disease resistance and PR protein production. 31 refs., 9 figs., 1 tab.

  15. Dataset on inflammatory proteins expressions and sialic acid levels in apolipoprotein E-deficient mice with administration of N-acetylneuraminic acid and/or quercetin.

    PubMed

    Dong, Rongrong; Li, Fahui; Qin, Shucun; Wang, Yi; Si, Yanhong; Xu, Xuelian; Tian, Hua; Zhai, Lei; Zhang, Guangjie; Li, Yongjun; Zhou, Yawei; Zhang, Ying; Zhang, Nan; Guo, Shoudong

    2016-09-01

    The data presented in this article describe an effect of N-acetylneuraminic acid and/or quercetin on the inflammatory proteins expressions (TNF-α, ICAM-1, VCAM-1 and MOMA-2) and the N-acetylneuraminic acid (NANA) levels of apolipoprotein E-deficient mice that are given a high-fat diet. Protein expression was performed by immunohistochemical imaging and NANA was quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) or semi-quantified using Image-Pro Plus software after ligation with fluorescein-5-thiosemicarbazide (FTSC). Further interpretation and discussion could be found at our research article entitled "Exogenous supplement of N-acetylneuraminic acid ameliorates atherosclerosis in apolipoprotein E-deficient mice" (Guo et al., 2016) [1]. PMID:27419199

  16. Signaling Pathways Related to Protein Synthesis and Amino Acid Concentration in Pig Skeletal Muscles Depend on the Dietary Protein Level, Genotype and Developmental Stages.

    PubMed

    Liu, Yingying; Li, Fengna; Kong, Xiangfeng; Tan, Bie; Li, Yinghui; Duan, Yehui; Blachier, François; Hu, Chien-An A; Yin, Yulong

    2015-01-01

    Muscle growth is regulated by the homeostatic balance of the biosynthesis and degradation of muscle proteins. To elucidate the molecular interactions among diet, pig genotype, and physiological stage, we examined the effect of dietary protein concentration, pig genotype, and physiological stages on amino acid (AA) pools, protein deposition, and related signaling pathways in different types of skeletal muscles. The study used 48 Landrace pigs and 48 pure-bred Bama mini-pigs assigned to each of 2 dietary treatments: lower/GB (Chinese conventional diet)- or higher/NRC (National Research Council)-protein diet. Diets were fed from 5 weeks of age to respective market weights of each genotype. Samples of biceps femoris muscle (BFM, type I) and longissimus dorsi muscle (LDM, type II) were collected at nursery, growing, and finishing phases according to the physiological stage of each genotype, to determine the AA concentrations, mRNA levels for growth-related genes in muscles, and protein abundances of mechanistic target of rapamycin (mTOR) signaling pathway. Our data showed that the concentrations of most AAs in LDM and BFM of pigs increased (P<0.05) gradually with increasing age. Bama mini-pigs had generally higher (P<0.05) muscle concentrations of flavor-related AA, including Met, Phe, Tyr, Pro, and Ser, compared with Landrace pigs. The mRNA levels for myogenic determining factor, myogenin, myocyte-specific enhancer binding factor 2 A, and myostatin of Bama mini-pigs were higher (P<0.05) than those of Landrace pigs, while total and phosphorylated protein levels for protein kinase B, mTOR, and p70 ribosomal protein S6 kinases (p70S6K), and ratios of p-mTOR/mTOR, p-AKT/AKT, and p-p70S6K/p70S6K were lower (P<0.05). There was a significant pig genotype-dependent effect of dietary protein on the levels for mTOR and p70S6K. When compared with the higher protein-NRC diet, the lower protein-GB diet increased (P<0.05) the levels for mTOR and p70S6K in Bama mini-pigs, but

  17. Common FABP4 Genetic Variants and Plasma Levels of Fatty Acid Binding Protein 4 in Older Adults

    PubMed Central

    Mukamal, Kenneth J.; Wilk, Jemma B.; Biggs, Mary L.; Jensen, Majken K.; Ix, Joachim H.; Kizer, Jorge R.; Tracy, Russell P.; Zieman, Susan J.; Mozaffarian, Dariush; Psaty, Bruce M.; Siscovick, David S.; Djoussé, Luc

    2013-01-01

    We examined common variants in the fatty acid binding protein 4 gene (FABP4) and plasma levels of FABP4 in adults aged 65 and older from the Cardiovascular Health Study. We genotyped rs16909187, rs1054135, rs16909192, rs10808846, rs7018409, rs2290201, and rs6992708 and measured circulating FABP4 levels among 3190 European Americans and 660 African Americans. Among European Americans, the minor alleles of six single nucleotide polymorphisms (SNP) were associated with lower FABP4 levels (all p ≤ 0.01). Among African Americans, the SNP with the lowest minor allele frequency was associated with lower FABP4 levels (p = 0.015). The C-A haplotype of rs16909192 and rs2290201 was associated with lower FABP4 levels in both European Americans (frequency = 16 %; p = 0.001) and African Americans (frequency = 8 %; p = 0.04). The haplotype combined a SNP in the first intron with one in the 3′untranslated region. However, the alleles associated with lower FABP4 levels were associated with higher fasting glucose in meta-analyses from the MAGIC consortium. These results demonstrate associations of common SNP and haplotypes in the FABP4 gene with lower plasma FABP4 but higher fasting glucose levels. PMID:24043587

  18. Effects of macromolecular crowding on a small lipid binding protein probed at the single-amino acid level.

    PubMed

    Pérez Santero, Silvia; Favretto, Filippo; Zanzoni, Serena; Chignola, Roberto; Assfalg, Michael; D'Onofrio, Mariapina

    2016-09-15

    Macromolecular crowding is a distinctive feature of the cellular interior, influencing the behaviour of biomacromolecules. Despite significant advancements in the description of the effects of crowding on global protein properties, the influence of cellular components on local protein attributes has received limited attention. Here, we describe a residue-level systematic interrogation of the structural, dynamic, and binding properties of the liver fatty acid binding protein (LFABP) in crowded solutions. Two-dimensional NMR spectral fingerprints and relaxation data were collected on LFABP in the presence of polymeric and biomolecular crowders. Non-interacting crowders produced minimal site-specific spectral perturbations on ligand-free and lipid-bound LFABP. Conformational adaptations upon ligand binding reproduced those observed in dilute solution, but a perturbation of the free oleate state resulted in less favorable uptake. When LFABP engaged in direct interactions with background molecules, changes in local chemical environments were detected for residues of the internal binding pocket and of the external surface. Enhanced complexity was introduced by investigating LFABP in cell lysates, and in membrane-bounded compartments. LFABP was able to capture ligands from prokaryotic and eukaryotic cell lysates, and from artificial cells (water-in-oil emulsion droplets). The data suggest that promiscuous interactions are a major factor influencing protein function in the cell. PMID:27457417

  19. Palmitic acid increase levels of pancreatic duodenal homeobox-1 and p38/stress-activated protein kinase in islets from rats maintained on a low protein diet.

    PubMed

    Arantes, Vanessa C; Reis, Marise A B; Latorraca, Márcia Q; Ferreira, Fabiano; Stoppiglia, Luiz Fabrízio; Carneiro, Everardo M; Boschero, Antonio C

    2006-12-01

    A severe reduction in insulin release in response to glucose is consistently noticed in protein-deprived rats and is attributed partly to the chronic exposure to elevated levels of NEFA. Since the pancreatic and duodenal transcription factor homeobox 1 (PDX-1) is important for the maintenance of beta-cell physiology, and since PDX-1 expression is altered in the islets of rats fed a low protein (LP) diet and that rats show high NEFA levels, we assessed PDX-1 and insulin mRNA expression, as well as PDX-1 and p38/stress activated protein kinase 2 (SAPK2) protein expression, in islets from young rats fed low (6%) or normal (17%; control) protein diets and maintained for 48 h in culture medium containing 5.6 mmol/l glucose, with or without 0.6 mmol/l palmitic acid. We also measured glucose-induced insulin secretion and glucose metabolism. Insulin secretion by isolated islets in response to 16.7 mmol/l glucose was reduced in LP compared with control rats. In the presence of NEFA, there was an increase in insulin secretion in both groups. At 2.8 mmol/l glucose, the metabolism of this sugar was reduced in LP islets, regardless of the presence of this fatty acid. However, when challenged with 16.7 mmol/l glucose, LP and control islets showed a severe reduction in glucose oxidation in the presence of NEFA. The PDX-1 and insulin mRNA were significantly higher when NEFA was added to the culture medium in both groups of islets. The effect of palmitic acid on PDX-1 and p38/SAPK2 protein levels was similar in LP and control islets, but the increase was much more evident in LP islets. These results demonstrate the complex interrelationship between nutrients in the control of insulin release and support the view that fatty acids play an important role in glucose homeostasis by affecting molecular mechanisms and stimulus/secretion coupling pathways. PMID:17181874

  20. Effect of temperature and concentration on benzoyl peroxide bleaching efficacy and benzoic acid levels in whey protein concentrate.

    PubMed

    Smith, T J; Gerard, P D; Drake, M A

    2015-11-01

    Much of the fluid whey produced in the United States is a by-product of Cheddar cheese manufacture and must be bleached. Benzoyl peroxide (BP) is currently 1 of only 2 legal chemical bleaching agents for fluid whey in the United States, but benzoic acid is an unavoidable by-product of BP bleaching. Benzoyl peroxide is typically a powder, but new liquid BP dispersions are available. A greater understanding of the bleaching characteristics of BP is necessary. The objective of the study was to compare norbixin destruction, residual benzoic acid, and flavor differences between liquid whey and 80% whey protein concentrates (WPC80) bleached at different temperatures with 2 different benzoyl peroxides (soluble and insoluble). Two experiments were conducted in this study. For experiment 1, 3 factors (temperature, bleach type, bleach concentration) were evaluated for norbixin destruction using a response surface model-central composite design in liquid whey. For experiment 2, norbixin concentration, residual benzoic acid, and flavor differences were explored in WPC80 from whey bleached by the 2 commercially available BP (soluble and insoluble) at 5 mg/kg. In liquid whey, soluble BP bleached more norbixin than insoluble BP, especially at lower concentrations (5 and 10 mg/kg) at both cold (4°C) and hot (50°C) temperatures. The WPC80 from liquid whey bleached with BP at 50°C had lower norbixin concentration, benzoic acid levels, cardboard flavor, and aldehyde levels than WPC80 from liquid whey bleached with BP at 4°C. Regardless of temperature, soluble BP destroyed more norbixin at lower concentrations than insoluble BP. The WPC80 from soluble-BP-bleached wheys had lower cardboard flavor and lower aldehyde levels than WPC80 from insoluble-BP-bleached whey. This study suggests that new, soluble (liquid) BP can be used at lower concentrations than insoluble BP to achieve equivalent bleaching and that less residual benzoic acid remains in WPC80 powder from liquid whey

  1. Effect of temperature and concentration on benzoyl peroxide bleaching efficacy and benzoic acid levels in whey protein concentrate.

    PubMed

    Smith, T J; Gerard, P D; Drake, M A

    2015-11-01

    Much of the fluid whey produced in the United States is a by-product of Cheddar cheese manufacture and must be bleached. Benzoyl peroxide (BP) is currently 1 of only 2 legal chemical bleaching agents for fluid whey in the United States, but benzoic acid is an unavoidable by-product of BP bleaching. Benzoyl peroxide is typically a powder, but new liquid BP dispersions are available. A greater understanding of the bleaching characteristics of BP is necessary. The objective of the study was to compare norbixin destruction, residual benzoic acid, and flavor differences between liquid whey and 80% whey protein concentrates (WPC80) bleached at different temperatures with 2 different benzoyl peroxides (soluble and insoluble). Two experiments were conducted in this study. For experiment 1, 3 factors (temperature, bleach type, bleach concentration) were evaluated for norbixin destruction using a response surface model-central composite design in liquid whey. For experiment 2, norbixin concentration, residual benzoic acid, and flavor differences were explored in WPC80 from whey bleached by the 2 commercially available BP (soluble and insoluble) at 5 mg/kg. In liquid whey, soluble BP bleached more norbixin than insoluble BP, especially at lower concentrations (5 and 10 mg/kg) at both cold (4°C) and hot (50°C) temperatures. The WPC80 from liquid whey bleached with BP at 50°C had lower norbixin concentration, benzoic acid levels, cardboard flavor, and aldehyde levels than WPC80 from liquid whey bleached with BP at 4°C. Regardless of temperature, soluble BP destroyed more norbixin at lower concentrations than insoluble BP. The WPC80 from soluble-BP-bleached wheys had lower cardboard flavor and lower aldehyde levels than WPC80 from insoluble-BP-bleached whey. This study suggests that new, soluble (liquid) BP can be used at lower concentrations than insoluble BP to achieve equivalent bleaching and that less residual benzoic acid remains in WPC80 powder from liquid whey

  2. Signaling Pathways Related to Protein Synthesis and Amino Acid Concentration in Pig Skeletal Muscles Depend on the Dietary Protein Level, Genotype and Developmental Stages

    PubMed Central

    Liu, Yingying; Li, Fengna; Kong, Xiangfeng; Tan, Bie; Li, Yinghui; Duan, Yehui; Blachier, François; Hu, Chien-An A.; Yin, Yulong

    2015-01-01

    Muscle growth is regulated by the homeostatic balance of the biosynthesis and degradation of muscle proteins. To elucidate the molecular interactions among diet, pig genotype, and physiological stage, we examined the effect of dietary protein concentration, pig genotype, and physiological stages on amino acid (AA) pools, protein deposition, and related signaling pathways in different types of skeletal muscles. The study used 48 Landrace pigs and 48 pure-bred Bama mini-pigs assigned to each of 2 dietary treatments: lower/GB (Chinese conventional diet)- or higher/NRC (National Research Council)-protein diet. Diets were fed from 5 weeks of age to respective market weights of each genotype. Samples of biceps femoris muscle (BFM, type I) and longissimus dorsi muscle (LDM, type II) were collected at nursery, growing, and finishing phases according to the physiological stage of each genotype, to determine the AA concentrations, mRNA levels for growth-related genes in muscles, and protein abundances of mechanistic target of rapamycin (mTOR) signaling pathway. Our data showed that the concentrations of most AAs in LDM and BFM of pigs increased (P<0.05) gradually with increasing age. Bama mini-pigs had generally higher (P<0.05) muscle concentrations of flavor-related AA, including Met, Phe, Tyr, Pro, and Ser, compared with Landrace pigs. The mRNA levels for myogenic determining factor, myogenin, myocyte-specific enhancer binding factor 2 A, and myostatin of Bama mini-pigs were higher (P<0.05) than those of Landrace pigs, while total and phosphorylated protein levels for protein kinase B, mTOR, and p70 ribosomal protein S6 kinases (p70S6K), and ratios of p-mTOR/mTOR, p-AKT/AKT, and p-p70S6K/p70S6K were lower (P<0.05). There was a significant pig genotype-dependent effect of dietary protein on the levels for mTOR and p70S6K. When compared with the higher protein-NRC diet, the lower protein-GB diet increased (P<0.05) the levels for mTOR and p70S6K in Bama mini-pigs, but

  3. The effect of N-stearoylethanolamine on cholesterol content, fatty acid composition and protein carbonylation level in rats with alimentary obesity-induced insulin resistance.

    PubMed

    Onopchenko, O V; Kosiakova, G V; Meged, E F; Klimashevsky, V M; Hula, N M

    2014-01-01

    The effect of N-stearoylethanolamine (NSE) on liver free fatty acid composition, cholesterol content and carbonylated protein level in rats with obesity-induced insulin resistance (IR) was studied in the work. The experimental insulin resistance was induced by prolonged high fat diet (58% of energy derived from fat) for 6 months combined with one injection of low-dose (15 mg/kg) of streptozotocin. The lipid assay showed a rise in liver free cholesterol content anda significant reduction in cholesterol esters level. Analyzing liver fatty acid composition, a decrease in polyunsaturated of fatty acid (PUFA) level and an increase in monounsaturated fatty acid (MUFA) content was found. Fatty acid imbalance with high content of MUFA was associated with elevated level ofprotein carbonylation. The NSE administration (50 mg/kg of body weight) for 2 weeks decreased free cholesterol content, increased cholesterol esters level and reduced free oleic fatty acid content in the liver of rats with IR. The effect of NSE on lipid imbalance led to a decrease in protein carbonylation level that may result in improvement of transmembrane protein function under obesity-induced insulin resistance state. PMID:25816612

  4. Estimation of endogenous protein and amino acid ileal losses in weaned piglets by regression analysis using diets with graded levels of casein

    PubMed Central

    2013-01-01

    Background Many studies have investigated endogenous loss of proteins and amino acids (AAs) at the ileal level in growing pigs. However, only a few studies have researched this subject in piglets. Knowledge regarding AA ileal digestibility in piglets would be helpful during the formulation of diets for weaning piglets, rather than just using coefficients obtained in growing pigs. Therefore, in this study, we sought to estimate endogenous protein and AA ileal losses in piglets. Furthermore, apparent and true ileal digestibility (AID and TID) of protein and AAs from casein were measured. Results The average flow of protein was 20.8 g/kg of dry matter intake (DMI). Basal protein loss, as estimated by regression, was 16.9 g/kg DMI. Glutamic acid, arginine, and aspartic acid (2.2, 1.4, and 1.2 g/kg DMI, respectively) were the AAs for which greater losses were seen. The AID of protein and AAs increased as the protein level in the diet increased. A higher increment in AID was observed between diets with 80 and160 g CP/kg of feed; this finding was mainly attributable to increases in glycine and arginine (46.1% and 18%, respectively). The TID of protein was 97.8, and the TID of AAs varied from 93.9 for histidine to 100.2 for phenylalanine. Conclusions The basal endogenous protein loss in piglets was 16.9 g/kg DMI. Endogenous protein was rich in glutamic acid, aspartic acid, and arginine, which represented 32.7% of endogenous protein loss in weaning piglets. The TID of casein was high and varied from 93.0 for histidine to 100.2 for phenylalanine. PMID:24053636

  5. Protein nutrition of Tenebrio molitor L. XX. Growth response of larvae to graded levels of amino acids.

    PubMed

    John, A M; Davis, G R; Sosulski, F W

    1979-12-01

    Larvae of Tenebrio molitor L;, Gembloux strain, race F, having an average initial weight of 10 mg were reared for 4 weeks at 27 +/- 0.25 degrees C and 65 +/- 5% relative humidity on diets containing an amino-acid mixture resembling the composition of larval tissues; Each of 14 amino acids was tested individually at levels of 0, 10, 20, 50, 100 and 200% of the amount found in larval tissues, while the concentration of other amino acids remained constant. Diets were maintained isonitrogenous at 1.6% nitrogen with supplementary glutamic acid, glycine, serine and tyrosine; Maximum fresh weight gains and dry weights were achieved by larvae fed reference levels of all essential amino acids except phenylalanine, threonine and tryptophan. Maximum growth was achieved by larvae fed 50% of the phenylalanine level found in larval tissues, Threonine and tryptophan were the limiting amino acids in this study and are probably required in the diet in excess of twice the concentration occuring in larval tissues, Probable ranges for quantitative amino-acid requirements of T. molitor were determined and suggestions were made for improving the nutritional adequacy of the amino-acid mixture.

  6. Apparent or Standardized Ileal Digestibility of Amino Acids of Diets Containing Different Protein Feedstuffs Fed at Two Crude Protein Levels for Growing Pigs

    PubMed Central

    Adebiyi, A. O.; Ragland, D.; Adeola, O.; Olukosi, O. A.

    2015-01-01

    The current study determined the apparent or standardized ileal digestibility of amino acids (AID or SID of AA) in growing pigs fed diets containing three protein feedstuffs with different fiber characteristics at two dietary crude protein (CP) levels. Twenty boars (Yorkshire×Landrace) with average initial body weight of 35 (±2.6) kg were fitted with a simple T-cannula at the distal ileum. These pigs were offered six diets containing soybean meal (SBM), canola meal (CM) or corn distillers dried grains with solubles (corn-DDGS) that were either adequate (19%) or marginal (15%) in CP using a triplicated 6×2 Youden Square Design. Except for Met, Trp, Cys, and Pro, AID of AA was greater (p<0.05) in the SBM diet compared with the CM diet. Apparent ileal digestibility for Gly and Asp was greater (p<0.05) in the SBM diet compared with the corn-DDGS diet. The AID of Ile, Leu, Phe, Val, Ala, Tyr, and Asp was greater (p<0.05) in the corn-DDGS diet compared with the CM diet. Standardized ileal digestibility of AA was greater (p<0.05) in the SBM diet compared with the CM diet for all AA except Trp and Pro. The SID of Ile, Leu, Val, Ala, Tyr, and Asp was greater (p<0.05) in the corn-DDGS diet compared with the CM diet. It was concluded that protein feedstuff affects ileal AA digestibility and is closely related to dietary fiber characteristics, and a 4-percentage unit reduction in dietary CP had no effect on ileal AA digestibility in growing pigs. PMID:26194226

  7. Overexpression of hydroxynitrile lyase in cassava roots elevates protein and free amino acids while reducing residual cyanogen levels.

    PubMed

    Narayanan, Narayanan N; Ihemere, Uzoma; Ellery, Claire; Sayre, Richard T

    2011-01-01

    Cassava is the major source of calories for more than 250 million Sub-Saharan Africans, however, it has the lowest protein-to-energy ratio of any major staple food crop in the world. A cassava-based diet provides less than 30% of the minimum daily requirement for protein. Moreover, both leaves and roots contain potentially toxic levels of cyanogenic glucosides. The major cyanogen in cassava is linamarin which is stored in the vacuole. Upon tissue disruption linamarin is deglycosylated by the apolplastic enzyme, linamarase, producing acetone cyanohydrin. Acetone cyanohydrin can spontaneously decompose at pHs >5.0 or temperatures >35°C, or is enzymatically broken down by hydroxynitrile lyase (HNL) to produce acetone and free cyanide which is then volatilized. Unlike leaves, cassava roots have little HNL activity. The lack of HNL activity in roots is associated with the accumulation of potentially toxic levels of acetone cyanohydrin in poorly processed roots. We hypothesized that the over-expression of HNL in cassava roots under the control of a root-specific, patatin promoter would not only accelerate cyanogenesis during food processing, resulting in a safer food product, but lead to increased root protein levels since HNL is sequestered in the cell wall. Transgenic lines expressing a patatin-driven HNL gene construct exhibited a 2-20 fold increase in relative HNL mRNA levels in roots when compared with wild type resulting in a threefold increase in total root protein in 7 month old plants. After food processing, HNL overexpressing lines had substantially reduced acetone cyanohydrin and cyanide levels in roots relative to wild-type roots. Furthermore, steady state linamarin levels in intact tissues were reduced by 80% in transgenic cassava roots. These results suggest that enhanced linamarin metabolism contributed to the elevated root protein levels.

  8. Effects of formula protein level and ratio on infant growth, plasma amino acids and serum trace elements. I. Cow's milk formula.

    PubMed

    Lönnerdal, B; Chen, C L

    1990-03-01

    The optimum level and ratios of protein to be used in cow's milk formula has recently been under discussion. Healthy term infants were fed from birth exclusively human milk or a formula that varied in protein level or whey: casein ratio: (A) 1.4 g/dl; 55:45, (B) 1.5 g/dl; 55:45, (C) 1.3 g/dl; 55:45, (D) 1.4 g/dl; 60:40, (E) 1.4 g/dl; 20:80. Infants were followed for 12 weeks and blood samples were taken at 2, 4, 8 and 12 weeks. Anthropometric indices did not show any significant differences among groups. Plasma amino acid and BUN levels of the C group were closest to the breast-fed group, while the formula with the highest protein level (B) resulted in high values for some amino acids. When comparing the formulas with 1.4 g protein/dl, the high casein group had the lowest plasma tryptophan levels. Taurine was added to all formulas at a level similar to that of breast milk; plasma taurine levels were similar for all groups. All formulas contained 0.7 mg iron and 0.7 mg zinc/dl; no differences were found among the groups in hematological indices or serum trace elements. These data show that feeding a formula with 1.3 g protein/dl and 55:45 whey: casein ratio from birth will result in growth and metabolic indices similar to those of breast-fed infants, although some plasma amino acid levels are not identical, 1990.

  9. Effect of pig breed and dietary protein level on selected fatty acids and stearoyl-coenzyme A desaturase protein expression in longissimus muscle and subcutaneous fat.

    PubMed

    Bessa, R J B; Hughes, R A; Jeronimo, E; Moreira, O C; Prates, J A M; Doran, O

    2013-09-01

    The aims of the study were 1) to investigate effects of a low protein diet on fatty acids content and composition of the LM and subcutaneous adipose tissue in 3 genetically diverse breeds, Large White × Landrace, Alentejano, and Bizaro, and 2) to determine whether the effect of the low protein diet of fatty acid composition is associated with dietary modulation of stearoyl-CoA desaturase (SCD) protein expression. The experiments were conducted on 12 Large White × Landrace, 12 Alentejano, and 10 Bízaro female and male pigs. The average animal BW at the beginning of experiments was 40.8, 40.7, and 38.3 kg for Large White × Landrace, Alentejano, and Bízaro, respectively, and the BW of animals at slaughter was 90 kg. The diets contained 202 or 169 g/kg DM of CP (high and low protein diets, respectively) and were balanced in essential AA. The diets were fed until the animals reached 90 kg BW (approximately 73 d). It was established that Large White × Landrace pigs had a less (P = 0.001) total fatty acid content in subcutaneous adipose tissue when compared with Alentejano and Bízaro and less (P < 0.001) intramuscular fat (IMF) content when compared with Alentejano. There was a positive relationship between SCD protein expression in the LM and MUFA content (r = 0.627, P = 0.029) and SCD protein expression and total muscle fatty acids content (r = 0.725, P = 0.008) in Large White × Landrace but not in Alentejano and Bizaro breeds. It has been suggested that SCD protein expression is associated with regulation of fat deposition only in breeds with genetic predisposition to a low IMF content.

  10. Accumulation of PrLeg, a Perilla legumin protein in potato tuber results in enhanced level of sulphur-containing amino acids.

    PubMed

    Goo, Young-Min; Kim, Tae-Won; Lee, Min-Kyung; Lee, Shin-Woo

    2013-09-01

    Potato is the fourth staple food in the world, following rice, wheat, and maize, whereas tubers contain high quality of starch, relatively high amounts of vitamin C and many other important substances. It also contains relatively good quality of protein (about 3 to 6% of the dried weight) and patatin, and 11S globulin is a major storage protein with high level of lysine. However, tuber protein contains relatively low amounts of sulphur-containing amino acids, which may result in low nutritional value. Recently, we cloned a gene encoding PrLeg polypeptide, a seed storage protein from Perilla, which contains relatively higher levels of sulphur-containing amino acids. We transformed PrLeg cDNA into a potato plant to over-express under the direction of the tuber-specific promoter, patatin. Most of the transgenic lines identified through PCR and RT-PCR analyses were able to accumulate high amount of prLeg transcript in their tuber tissue, while very little or no transcript that were detected in their leaf tissues. The level of methionine content was elevated up to three-fold compared to non-transgenic parental line, without any significant changes in other amino acids, suggesting that further research is required to get a deeper insight into their nutritional value. PMID:24161240

  11. Serum copper, ceruloplasmin, protein thiols and thiobarbituric acid reactive substance status in liver cancer associated with elevated levels of alpha-fetoprotein.

    PubMed

    Nayak, Shivananda B; Yashwanth, S; Pinto, Sneha M; Bhat, Vinutha R; Mayya, Srimathi S

    2005-01-01

    Serum copper, ceruloplasmin, protein thiols and thiobarbituric acid reactive substances (TEARS) were estimated in 25 patients of liver cancer. The copper to ceruloplasmin ratio was moderately increased (P<0.05) but the copper (P<0.001) and ceruloplasmin (P<0.001) levels were significantly increased in liver cancer patients when compared to controls. Protein thiols levels were found to be highly significant (P<0.001). Where as the TEARS levels were not found to be significant. Trace elements and free radicals have been implicated in the etiology of cancer. Hence the estimation of ceruloplasmin and protein thiols along with the copper may be of high value in the early diagnosis of cancer.

  12. A fish protein hydrolysate alters fatty acid composition in liver and adipose tissue and increases plasma carnitine levels in a mouse model of chronic inflammation

    PubMed Central

    2013-01-01

    Background There is growing evidence that fish protein hydrolysate (FPH) diets affect mitochondrial fatty acid metabolism in animals. The aim of the study was to determine if FPH could influence fatty acid metabolism and inflammation in transgene mice expressing human tumor necrosis factor alpha (hTNFα). Methods hTNFα mice (C57BL/6 hTNFα) were given a high-fat (23%, w/w) diet containing 20% casein (control group) or 15% FPH and 5% casein (FPH group) for two weeks. After an overnight fast, blood, adipose tissue, and liver samples were collected. Gene expression and enzyme activity was analysed in liver, fatty acid composition was analyzed in liver and ovarian white adipose tissue, and inflammatory parameters, carnitine, and acylcarnitines were analyzed in plasma. Results The n-3/n-6 fatty acid ratio was higher in mice fed the FPH diet than in mice fed the control diet in both adipose tissue and liver, and the FPH diet affected the gene expression of ∆6 and ∆9 desaturases. Mice fed this diet also demonstrated lower hepatic activity of fatty acid synthase. Concomitantly, a lower plasma INF-γ level was observed. Plasma carnitine and the carnitine precursor γ-butyrobetaine was higher in the FPH-group compared to control, as was plasma short-chained and medium-chained acylcarnitine esters. The higher level of plasma acetylcarnitine may reflect a stimulated mitochondrial and peroxisomal β-oxidation of fatty acids, as the hepatic activities of peroxisomal acyl-CoA oxidase 1 and mitochondrial carnitine palmitoyltransferase-II were higher in the FPH-fed mice. Conclusions The FPH diet was shown to influence hepatic fatty acid metabolism and fatty acid composition. This indicates that effects on fatty acid metabolism are important for the bioactivity of protein hydrolysates of marine origin. PMID:24098955

  13. Folic acid and protein content in maternal diet and postnatal high-fat feeding affect the tissue levels of iron, zinc, and copper in the rat.

    PubMed

    Król, Ewelina; Krejpcio, Zbigniew; Chmurzynska, Agata

    2011-12-01

    Although maternal, fetal, and placental mechanisms compensate for disturbances in the fetal environment, any nutritional inadequacies present during pregnancy may affect fetal metabolism, and their consequences may appear in later life. The aim of the present study is to investigate the influence of maternal diet during gestation on Fe, Zn, and Cu levels in the livers and kidneys of adult rats. The study was carried out on the offspring (n = 48) of mothers fed either a protein-balanced or a protein-restricted diet (18% vs. 9% casein) during pregnancy, with or without folic acid supplementation (0.005- vs. 0.002-g folic acid/kg diet). At 10 weeks of age, the offspring of each maternal group were randomly assigned to groups fed either the AIN-93G diet or a high-fat diet for 6 weeks, until the end of the experiment. The levels of Fe, Zn, and Cu in the livers and kidneys were determined by the F-AAS method. It was found that postnatal exposure to the high-fat diet was associated with increased hepatic Fe levels (p < 0.001), and with decreased liver Zn and Cu contents (p < 0.01 and p < 0.05, respectively), as well as with decreased renal Cu contents (p < 0.001). Moreover, the offspring's tissue mineral levels were also affected by protein and folic acid content in the maternal diet. Both prenatal protein restriction and folic acid supplementation increased the liver Zn content (p < 0.05) and the kidney Zn content (p < 0.001; p < 0.05, respectively), while folic acid supplementation resulted in a reduction in renal Cu level (p < 0.05). Summarizing, the results of this study show that maternal dietary folic acid and protein intake during pregnancy, as well as the type of postweaning diet, affect Fe, Zn, and Cu levels in the offspring of the rat. However, the mechanisms responsible for this phenomenon are unclear, and warrant further investigation.

  14. Regulation of insulin-like growth factor-binding protein messenger ribonucleic acid levels in sheep thyroid cells.

    PubMed

    Bachrach, L K; Eggo, M C; Burrow, G N; Liu, F; Tram, T; Powell, D R

    1991-04-01

    The insulin-like growth factors (IGFs) exist primarily bound to cell surface receptors or complexed to specific binding proteins (IGFBPs). The IGFBPs modulate the bioavailability of the IGFs and may enhance or inhibit IGF actions. Several distinct forms of IGFBPs have been described on the basis of size, immunological determinants, and distribution in biological fluids; the IGFBPs may differ as well in their biological function. Sheep thyroid cells produce IGFBPs under hormonal regulation. Cells grown in basal medium or with six-hormone (6H) medium supplements (transferrin, glycyl-histidyl-lysine, hydrocortisone, somatostatin, insulin, and TSH) release nonglycosylated BPs that migrate at 24, 27, 29, and 32 kDa on Western ligand blot. Cells cultured with the thyroid mitogens epidermal growth factor and phorbol ester release additional glycosylated IGFBPs of 40-44 kDa. Immunoprecipitation experiments indicate that 29- and 32-kDa IGFBPs are antigenically related to IGFBP-2, and the 40- to 44-kDa proteins are related to IGFBP-3. Using specific cDNA probes IGFBP-1, -2, and -3, we examined the regulation of IGFBP mRNA levels in sheep thyroid cultures. The rat IGFBP-2 cDNA probe hybridized to an approximately 1.6-kilobase mRNA species in cells under all culture conditions. However, IGFBP-3 mRNA was detectable only in epidermal growth factor- or phorbol ester-treated cells and appeared within 4 h, preceding the release of IGFBP-3 protein into the medium. The 6H additives, which stimulate differentiated function in thyroid cells, inhibited the mRNA levels of both IGFBP-2 and IGFBP-3. IGFBP-1 mRNA was not detectable. The distinct regulation of these IGFBPs suggest that they may play different biological roles in modulating thyroid physiology. PMID:1706262

  15. The Prognostic Value of Serum Levels of Heart-Type Fatty Acid Binding Protein and High Sensitivity C-Reactive Protein in Patients With Increased Levels of Amino-Terminal Pro-B Type Natriuretic Peptide

    PubMed Central

    Jeong, Ji Hun; Seo, Yiel Hea; Ahn, Jeong Yeal; Kim, Kyung Hee; Seo, Ja Young; Kim, Moon Jin; Lee, Hwan Tae

    2016-01-01

    Background Amino-terminal pro-B type natriuretic peptide (NT-proBNP) is a well-established prognostic factor in heart failure (HF). However, numerous causes may lead to elevations in NT-proBNP, and thus, an increased NT-proBNP level alone is not sufficient to predict outcome. The aim of this study was to evaluate the utility of two acute response markers, high sensitivity C-reactive protein (hsCRP) and heart-type fatty acid binding protein (H-FABP), in patients with an increased NT-proBNP level. Methods The 278 patients were classified into three groups by etiology: 1) acute coronary syndrome (ACS) (n=62), 2) non-ACS cardiac disease (n=156), and 3) infectious disease (n=60). Survival was determined on day 1, 7, 14, 21, 28, 60, 90, 120, and 150 after enrollment. Results H-FABP (P<0.001), NT-proBNP (P=0.006), hsCRP (P<0.001) levels, and survival (P<0.001) were significantly different in the three disease groups. Patients were divided into three classes by using receiver operating characteristic curves for NT-proBNP, H-FABP, and hsCRP. Patients with elevated NT-proBNP (≥3,856 pg/mL) and H-FABP (≥8.8 ng/mL) levels were associated with higher hazard ratio for mortality (5.15 in NT-proBNP and 3.25 in H-FABP). Area under the receiver operating characteristic curve analysis showed H-FABP was a better predictor of 60-day mortality than NT-proBNP. Conclusions The combined measurement of H-FABP with NT-proBNP provides a highly reliable means of short-term mortality prediction for patients hospitalized for ACS, non-ACS cardiac disease, or infectious disease. PMID:27374706

  16. Intake of branched-chain amino acids influences the levels of MAFbx mRNA and MuRF-1 total protein in resting and exercising human muscle.

    PubMed

    Borgenvik, Marcus; Apró, William; Blomstrand, Eva

    2012-03-01

    Resistance exercise and amino acids are two major factors that influence muscle protein turnover. Here, we examined the effects of resistance exercise and branched-chain amino acids (BCAA), individually and in combination, on the expression of anabolic and catabolic genes in human skeletal muscle. Seven subjects performed two sessions of unilateral leg press exercise with randomized supplementation with BCAA or flavored water. Biopsies were collected from the vastus lateralis muscle of both the resting and exercising legs before and repeatedly after exercise to determine levels of mRNA, protein phosphorylation, and amino acid concentrations. Intake of BCAA reduced (P < 0.05) MAFbx mRNA by 30 and 50% in the resting and exercising legs, respectively. The level of MuRF-1 mRNA was elevated (P < 0.05) in the exercising leg two- and threefold under the placebo and BCAA conditions, respectively, whereas MuRF-1 total protein increased by 20% (P < 0.05) only in the placebo condition. Phosphorylation of p70(S6k) increased to a larger extent (∼2-fold; P < 0.05) in the early recovery period with BCAA supplementation, whereas the expression of genes regulating mTOR activity was not influenced by BCAA. Muscle levels of phenylalanine and tyrosine were reduced (13-17%) throughout recovery (P < 0.05) in the placebo condition and to a greater extent (32-43%; P < 0.05) following BCAA supplementation in both resting and exercising muscle. In conclusion, BCAA ingestion reduced MAFbx mRNA and prevented the exercise-induced increase in MuRF-1 total protein in both resting and exercising leg. Further-more, resistance exercise differently influenced MAFbx and MuRF-1 mRNA expression, suggesting both common and divergent regulation of these two ubiquitin ligases.

  17. Proteomic investigation of protein profile changes and amino acid residue-level modification in cooked lamb longissimus thoracis et lumborum: The effect of roasting.

    PubMed

    Yu, Tzer-Yang; Morton, James D; Clerens, Stefan; Dyer, Jolon M

    2016-09-01

    Protein modifications of meat cooked by typical dry-heat methods (e.g., roasting) are currently not well understood. The present study utilised a shotgun proteomic approach to examine the molecular-level effect of roasting on thin lamb longissimus thoracis et lumborum patties, in terms of changes to both the protein profile and amino acid residue side-chain modifications. Cooking caused aggregation of actin, myosin heavy chains and sarcoplasmic proteins. Longer roasting time resulted in significantly reduced protein extractability as well as protein truncation involving particularly a number of myofibrillar and sarcoplasmic proteins, e.g., 6-phosphofructokinase, beta-enolase, l-lactate dehydrogenase A chain, alpha-actinin-3, actin and possibly myosin heavy chains. Modifications that have potential influence on nutritional properties, including carboxyethyllysine and a potentially glucose-derived N-terminal Amadori compound, were observed in actin and myoglobin after roasting. This study provided new insights into molecular changes resulting from the dry-heat treatment of meat, such as commonly used in food preparation. PMID:27150797

  18. Influence of liposomes rich in unsaturated or saturated fatty acids on the growth of human xenotransplanted mammary carcinomas and on the levels of heart type fatty acid binding protein.

    PubMed

    Naundorf, H; Zschiesche, W; Reszka, R; Fichtner, I

    1995-01-01

    A panel of 4 human mammary carcinomas passaged in nude mice were subjected to intraperitoneal application of cholesterol-free liposomes enriched with linoleic (unsaturated fatty acid) or stearic acid (saturated fatty acid). The liposomes were examined with regard to their influence on the tumor growth and level of heart type fatty acid binding protein (FABP). Liposomes with different fatty acid composition influenced the growth of mammary carcinomas 3366, BO, 4000 and 4151 in distinct ways. Liposomes with a high content of stearic acid significantly inhibited the growth of mammary carcinomas 3366 and BO, whereas mammary carcinomas 4000 and 4151 were not affected. The growth of mammary carcinoma 3366 was moderately increased after supplementation of liposomes rich in linoleic acid, the tumor BO was significantly inhibited and the growth of MaCa 4000 and 4151 was unchanged. Liposome treatment led to a significant increase in heart type FABP in mammary carcinomas 3366 and BO regardless of whether the animals were treated with liposomes rich in stearic or linoleic acid. Such significant changes of FABP level could not be observed in mammary carcinomas 4000 or 4151. We suggest that the lipid-mediated growth modulation seems to be dependent on an increase of heart type FABPs in these tumor models. PMID:8562891

  19. Effects of prenatal exposure to 2,4-D/2,4,5-T mixture on postnatal changes in rat brain glutamate, GABA protein, and nucleic acid levels

    SciTech Connect

    Mohammad, F.K.; Omer, V.E.V.

    1988-02-01

    The opportunity of maternal exposure to various chemicals in the work place and the general environments have increased, and the fetus and neonate may be at greater risk than the adult. However, the embryotoxic and teratogenic effects of the chlorinated phenoxy herbicides 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), the main chemicals in Agent Orange, are well documented only in laboratory animals. The brain of the developing fetus is vulnerable to the toxic effects of the phenoxy herbicides which readily cross the placental barrier and distribute into fetal tissues, including brain. Although the neurochemical basis for the behavioral teratogenicity of the phenoxy herbicides is not know, it was recently reported that non-teratogenic doses of a 1:1 mixture of 2,4-D and 2,4,5-T delayed the ontogeny of dopamine and serotonin in the brain of the developing rate. This communication provides further descriptive information about the ontogeny of rat brain nucleic acid, protein, glutamate and ..gamma..-aminobutyrate (GABA) following in utero exposure to non-teratogenic levels of a 1:1 mixture of 2,4-D/2,4,5-T.

  20. Sex in Drosophila mauritiana: a very high level of amino acid polymorphism in a male reproductive protein gene, Acp26Aa.

    PubMed

    Tsaur, S C; Ting, C T; Wu, C I

    2001-01-01

    Many genes pertaining to male reproductive functions have been shown to evolve rapidly between species, and evidence increasingly suggest the influence of positive Darwinian selection. The accessory gland protein gene (Acp26Aa) of Drosophila is one such example. In order to understand the mechanism of selection, it is often helpful to examine the pattern of polymorphism. We report here that the level of amino acid polymorphism in the N-terminal quarter of Acp26Aa is high in Drosophila melanogaster and is unprecedented in its sibling species Drosophila mauritiana. We postulate that (1) this N-terminal segment may play a role in sperm competition, and (2) D. mauritiana may have been under much more intense sexual selection than other species. Both postulates have important ramifications and deserve to be tested rigorously.

  1. Serum Level of Heart-Type Fatty Acid Binding Protein (H-FABP) Before and After Treatment of Congestive Heart Failure in Children.

    PubMed

    Zoair, Amr; Mawlana, Wegdan; Abo-Elenin, Amany; Korrat, Mostafa

    2015-12-01

    Remodeling of the heart following injury affects the morbidity and mortality in children presented with heart failure (HF). Heart-type fatty acid binding protein (H-FABP) is a novel biomarker that could be of help to predict the prognosis and risk stratification in those children. We aimed to evaluate the diagnostic and prognostic value of H-FABP in children with heart failure before and after treatment. The study was conducted as a prospective cohort study. It included 30 children with HF as a patient group and 20 healthy children matched for age and sex as a control group. Echocardiographic assessment of the heart was done using conventional Doppler echocardiography. Serum levels of (H-FABP) were measured using enzyme-linked immunosorbent assay before and after treatment of HF. All patients were observed during follow-up period of 3 months. There was a significant difference in the serum level of H-FABP in our patients before treatment (5.278 ± 3.253 ng/ml) compared with after treatment (2.089 ± 0.160 ng/ml) with significant difference compared with the control group. There was a significant increase in the serum level of H-FABP with increase in the severity of heart failure according to Ross classification. Significant increase in the H-FABP was associated with adverse outcome. Serum levels of H-FABP strongly correlated with clinical and echocardiographic assessment of LV performance of children with HF, and its levels significantly increased in children with adverse outcome suggesting its value as a useful diagnostic and prognostic predictor (with high sensitivity and specificity). PMID:26123812

  2. Effects of high peanut meal with different crude protein level supplemented with amino acids on performance, carcass traits and nitrogen retention of Chinese Yellow broilers.

    PubMed

    Gou, Z Y; Jiang, S Q; Jiang, Z Y; Zheng, C T; Li, L; Ruan, D; Chen, F; Lin, X J

    2016-08-01

    This study assessed the effects of feeding high peanut meal diets of reduced crude protein (CP) content supplemented with essential amino acids (EAA) on growth performance, carcass traits, biochemical indices in plasma, and nitrogen (N) retention of male and female Lingnan Yellow broilers from day 22 to day 42 of age. Each of four dietary treatments (19%, 18%, 17% or 16% CP, dietary CP level reduced by the reduced dietary peanut meal) contained six replicate pens with 35 birds of each sex (males and females with equal number), separately (1680 in total). The three diets with reduced CP were supplemented with 5 EAA to meet the requirements and provide the same levels as in the 19% CP diet. Average daily gain decreased and feed:gain ratio was worse in both sexes with reduced CP% (linear, p < 0.05). Dressing percentage increased as CP% decreased in males (linear, p < 0.05) and thigh muscle percentage reduced slightly in females (linear, p < 0.05). Abdominal fat percentage of males fed the 17% CP was the lowest (quadratic, p < 0.05). The plasma metabolic indices, concentrations of triglycerides and malondialdehyde, showed linear responses to reduced CP% (p < 0.05) with triglycerides increasing while malondialdehyde decreased. Plasma uric acid increased in females (linear, p < 0.05), but not in males, as CP% decreased. Efficiency of N retention increased and N excretion strikingly decreased with lower CP diets (p < 0.001), and both variables showed significant (p < 0.05) linear and quadratic effects. It is concluded that there was a limit to which dietary CP of broilers could be reduced without adverse effects. Dietary CP could be reduced to 17% for males and 18% for females (or 18% when fed together) between day 22 and day 42, if diets are supplemented with synthetic EAA. PMID:27401885

  3. Increased adenosine levels in mice expressing mutant glial fibrillary acidic protein in astrocytes result in failure of induction of LTP reversal (depotentiation) in hippocampal CA1 neurons.

    PubMed

    Fujii, Satoshi; Tanaka, Kenji F; Ikenaka, Kazuhiro; Yamazaki, Yoshihiko

    2014-08-26

    Astrocytes regulate the activity of neighboring neurons by releasing chemical transmitters, including ATP. Adenosine levels in the cerebrospinal fluid of mice that express a mutant human glial fibrillary acidic protein in astrocytes are slightly elevated compared to those in wild type mice and this might result from the observed increased release by mutant astrocytes of ATP, which can be used to produce adenosine. Using hippocampal slices from these mutant mice, we examined whether the increased endogenous adenosine levels in the hippocampus modulate the reversal of long-term potentiation (LTP), i.e. depotentiation (DP), in CA1 neurons. In hippocampal slices from wild type mice, a stable LTP was induced by tetanic stimulation consisting of 100 pulses at 100 Hz, and this was reversed by a train of low frequency stimulation (LFS) of 500 pulses at 1 Hz applied 30 min later. This induction of DP was inhibited by application of either 100 nM adenosine or 0.5 nM N(6)-cyclopentyladenosine, an adenosine A1 receptor agonist, during LFS, indicating that the increase in extracellular adenosine levels attenuated DP induction by acting on adenosine A1 receptors. In contrast, although a stable LTP was also induced in hippocampal slices from mutant mice, induction of DP was inhibited, but DP could be induced by application, during LFS, of 50 nM 8-cyclopentyltheophylline, an adenosine A1 receptor antagonist. These results suggest that a small increase in extracellular adenosine levels resulting from increased ATP release by astrocytes results in attenuation of DP in hippocampal CA1 neurons in the mutant mice.

  4. Increased adenosine levels in mice expressing mutant glial fibrillary acidic protein in astrocytes result in failure of induction of LTP reversal (depotentiation) in hippocampal CA1 neurons.

    PubMed

    Fujii, Satoshi; Tanaka, Kenji F; Ikenaka, Kazuhiro; Yamazaki, Yoshihiko

    2014-08-26

    Astrocytes regulate the activity of neighboring neurons by releasing chemical transmitters, including ATP. Adenosine levels in the cerebrospinal fluid of mice that express a mutant human glial fibrillary acidic protein in astrocytes are slightly elevated compared to those in wild type mice and this might result from the observed increased release by mutant astrocytes of ATP, which can be used to produce adenosine. Using hippocampal slices from these mutant mice, we examined whether the increased endogenous adenosine levels in the hippocampus modulate the reversal of long-term potentiation (LTP), i.e. depotentiation (DP), in CA1 neurons. In hippocampal slices from wild type mice, a stable LTP was induced by tetanic stimulation consisting of 100 pulses at 100 Hz, and this was reversed by a train of low frequency stimulation (LFS) of 500 pulses at 1 Hz applied 30 min later. This induction of DP was inhibited by application of either 100 nM adenosine or 0.5 nM N(6)-cyclopentyladenosine, an adenosine A1 receptor agonist, during LFS, indicating that the increase in extracellular adenosine levels attenuated DP induction by acting on adenosine A1 receptors. In contrast, although a stable LTP was also induced in hippocampal slices from mutant mice, induction of DP was inhibited, but DP could be induced by application, during LFS, of 50 nM 8-cyclopentyltheophylline, an adenosine A1 receptor antagonist. These results suggest that a small increase in extracellular adenosine levels resulting from increased ATP release by astrocytes results in attenuation of DP in hippocampal CA1 neurons in the mutant mice. PMID:25017946

  5. Proteins and acids from petroleum.

    PubMed

    Zaki, D; el-Badrawy, S

    1978-01-01

    The wax distillate fraction (boiling range 300 up to 400 degrees C) from the crude oil "El-Alameen" was found to be a good substrate for the biosynthesis of proteins and/or amino acids by bacteria under special culture conditions. The fermentation processes were accompanied by a refining effect to the oil fraction, elevating its refraction index and lowering its melting point, giving dewaxing effect to the oil fraction. PMID:735504

  6. Rosuvastatin Decreases Intestinal Fatty Acid Binding Protein (I-FABP), but Does Not Alter Zonulin or Lipopolysaccharide Binding Protein (LBP) Levels, in HIV-Infected Subjects on Antiretroviral Therapy

    PubMed Central

    Funderburg, Nicholas T.; Boucher, Morgan; Sattar, Abdus; Kulkarni, Manjusha; Labbato, Danielle; Kinley, Bruce I.; McComsey, Grace A.

    2016-01-01

    Introduction Altered gastrointestinal (GI) barrier integrity and subsequent microbial translocation may contribute to immune activation in HIV infection. We have reported that rosuvastatin improved several markers of immune activation in HIV+ participants, but the effect of statin treatment on markers of GI barrier dysfunction is unknown. Methods SATURN-HIV is a randomized, double-blind, placebo-controlled trial assessing the effect of rosuvastatin (10mg/daily) on markers of cardiovascular disease, inflammation, and immune activation in ART-treated patients. Gut-barrier integrity was assessed by the surrogate markers intestinal fatty acid binding protein (I-FABP), a marker of enterocyte death, and zonulin-1, a marker of gut epithelial cell function. Levels of lipopolysaccharide binding protein (LBP) were measured as a marker of microbial translocation. Results Rosuvastatin significantly reduced levels of I-FABP during the treatment period compared to the placebo. There was no effect of rosuvastatin treatment on levels of zonulin or LBP. Baseline levels of LBP were directly related to several markers of immune activation in samples from all participants, including soluble CD163, IP-10, VCAM-1, TNFR-II, and the proportion of CD4+ and CD8+ T cells expressing CD38 and HLA-DR. Many of these relationships, however, were not seen in the statin arm alone at baseline or over time, as inflammatory markers often decreased and LBP levels were unchanged. Conclusions Forty-eight weeks of rosuvastatin treatment reduced levels of I-FABP, but did not affect levels of zonulin or LBP. The reduction in levels of inflammatory markers that we have reported with rosuvastatin treatment is likely mediated through other mechanisms not related to gut integrity or microbial translocation. PMID:27500282

  7. CypD−/− Hearts HaveAltered Levels of Proteins Involved in Krebs Cycle, Branch Chain Amino Acid Degradation and Pyruvate Metabolism

    PubMed Central

    Menazza, Sara; Wong, Renee; Nguyen, Tiffany; Wang, Guanghui; Gucek, Marjan; Murphy, Elizabeth

    2013-01-01

    Cyclophilin D (CypD) is a mitochondrial chaperone that has been shown to regulate the mitochondrial permeability transition pore (MPTP). MPTP opening is a major determinant of mitochondrial dysfunction and cardiomyocyte death during ischemia/reperfusion (I/R) injury. Mice lacking CypD have been widely used to study regulation of the MPTP, and it has been shown recently that genetic depletion of CypD correlates with elevated levels of mitochondrial Ca2+. The present study aimed to characterize the metabolic changes in CypD−/− hearts. Initially, we used a proteomics approach to examine protein changes in CypD−/− mice. Using pathway analysis we found that CypD−/− hearts have alteration in branched chain amino acid metabolism, pyruvate metabolism and the Krebs cycle. We tested whether these metabolic changes were due to inhibition of electron transfer from these metabolic pathways into the electron transport chain. As we found decreased levels of succinate dehydrogenase and electron transfer flavoprotein in the proteomics analysis, we examined whether activities of these enzymes might be altered. However, we found no alterations in their activities. The proteomics study also showed a 23% decrease in carnitine -palmitoyltransferase 1 (CPT1), which prompted us to perform a metabolomics analysis. Consistent with the decrease in CPT1, we found a significant decrease in C4/Ci4, C5-OH/C3-DC, C12:1, C14:1, C16:1, and C20:3 acyl carnitines in hearts from CypD−/− mice. In summary, CypD−/− hearts exhibit changes in many metabolic pathways and caution should be used when interpreting results from these mice as due solely to inhibition of the MPTP. PMID:23262437

  8. CypD(-/-) hearts have altered levels of proteins involved in Krebs cycle, branch chain amino acid degradation and pyruvate metabolism.

    PubMed

    Menazza, Sara; Wong, Renee; Nguyen, Tiffany; Wang, Guanghui; Gucek, Marjan; Murphy, Elizabeth

    2013-03-01

    Cyclophilin D (CypD) is a mitochondrial chaperone that has been shown to regulate the mitochondrial permeability transition pore (MPTP). MPTP opening is a major determinant of mitochondrial dysfunction and cardiomyocyte death during ischemia/reperfusion (I/R) injury. Mice lacking CypD have been widely used to study regulation of the MPTP, and it has been shown recently that genetic depletion of CypD correlates with elevated levels of mitochondrial Ca(2+). The present study aimed to characterize the metabolic changes in CypD(-/-) hearts. Initially, we used a proteomics approach to examine protein changes in CypD(-/-) mice. Using pathway analysis, we found that CypD(-/-) hearts have alterations in branched chain amino acid metabolism, pyruvate metabolism and the Krebs cycle. We tested whether these metabolic changes were due to inhibition of electron transfer from these metabolic pathways into the electron transport chain. As we found decreased levels of succinate dehydrogenase and electron transfer flavoprotein in the proteomics analysis, we examined whether activities of these enzymes might be altered. However, we found no alterations in their activities. The proteomics study also showed a 23% decrease in carnitine-palmitoyltransferase 1 (CPT1), which prompted us to perform a metabolomics analysis. Consistent with the decrease in CPT1, we found a significant decrease in C4/Ci4, C5-OH/C3-DC, C12:1, C14:1, C16:1, and C20:3 acyl carnitines in hearts from CypD(-/-) mice. In summary, CypD(-/-) hearts exhibit changes in many metabolic pathways and caution should be used when interpreting results from these mice as due solely to inhibition of the MPTP.

  9. Dietary gamma-linolenic acid in the form of borage oil causes less body fat accumulation accompanying an increase in uncoupling protein 1 mRNA level in brown adipose tissue.

    PubMed

    Takahashi, Y; Ide, T; Fujita, H

    2000-10-01

    Rats were fed a low-fat diet containing 2% safflower oil or 20% fat diets containing either safflower oil rich in linoleic acid, borage oil containing 25% gamma (gamma)-linolenic acid or enzymatically prepared gamma-linolenic acid enriched borage oil containing 47% gamma-linolenic acid for 14 days. Energy intake and growth of animals were the same among groups. A high safflower oil diet compared with a low-fat diet caused significant increases in both epididymal and perirenal white adipose tissue weights. However, high-fat diets rich in gamma-linolenic acid failed to do so. Compared with a low-fat diet, all the high-fat diets increased mRNA levels of uncoupling protein 1 and lipoprotein lipase in brown adipose tissue. The extents of the increase were greater with high-fat diets rich in gamma-linolenic acid. Various high-fat diets, compared with a low-fat diet, decreased glucose transporter 4 mRNA in white adipose tissue to the same levels. The amount and types of dietary fat did not affect the leptin mRNA level in epididymal white adipose tissue. However, a high safflower oil diet, but not high-fat diets rich in gamma-linolenic acid relative to a low-fat diet, increased perirenal white adipose tissue leptin mRNA levels. All high-fat diets, relative to a low-fat diet, increased the hepatic mitochondrial fatty acid oxidation rate and fatty acid oxidation enzyme mRNA abundances to the same levels. High-fat diets also increased these parameters in the peroxisomal pathway, and the increases were greater with high-fat diets rich in gamma-linolenic acid. The physiological activity in increasing brown adipose tissue gene expression and peroxisomal fatty acid oxidation was similar between the two types of borage oil differing in gamma-linolenic acid content. It was suggested that dietary gamma-linolenic acid attenuates body fat accumulation through the increase in gene expressions of uncoupling protein 1 in brown adipose tissue. An increase in hepatic peroxisomal fatty acid

  10. Dietary gamma-linolenic acid in the form of borage oil causes less body fat accumulation accompanying an increase in uncoupling protein 1 mRNA level in brown adipose tissue.

    PubMed

    Takahashi, Y; Ide, T; Fujita, H

    2000-10-01

    Rats were fed a low-fat diet containing 2% safflower oil or 20% fat diets containing either safflower oil rich in linoleic acid, borage oil containing 25% gamma (gamma)-linolenic acid or enzymatically prepared gamma-linolenic acid enriched borage oil containing 47% gamma-linolenic acid for 14 days. Energy intake and growth of animals were the same among groups. A high safflower oil diet compared with a low-fat diet caused significant increases in both epididymal and perirenal white adipose tissue weights. However, high-fat diets rich in gamma-linolenic acid failed to do so. Compared with a low-fat diet, all the high-fat diets increased mRNA levels of uncoupling protein 1 and lipoprotein lipase in brown adipose tissue. The extents of the increase were greater with high-fat diets rich in gamma-linolenic acid. Various high-fat diets, compared with a low-fat diet, decreased glucose transporter 4 mRNA in white adipose tissue to the same levels. The amount and types of dietary fat did not affect the leptin mRNA level in epididymal white adipose tissue. However, a high safflower oil diet, but not high-fat diets rich in gamma-linolenic acid relative to a low-fat diet, increased perirenal white adipose tissue leptin mRNA levels. All high-fat diets, relative to a low-fat diet, increased the hepatic mitochondrial fatty acid oxidation rate and fatty acid oxidation enzyme mRNA abundances to the same levels. High-fat diets also increased these parameters in the peroxisomal pathway, and the increases were greater with high-fat diets rich in gamma-linolenic acid. The physiological activity in increasing brown adipose tissue gene expression and peroxisomal fatty acid oxidation was similar between the two types of borage oil differing in gamma-linolenic acid content. It was suggested that dietary gamma-linolenic acid attenuates body fat accumulation through the increase in gene expressions of uncoupling protein 1 in brown adipose tissue. An increase in hepatic peroxisomal fatty acid

  11. Oleic acid-dependent modulation of Nitric oxide associated 1 protein levels regulates nitric oxide-mediated defense signaling in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The conserved cellular metabolites nitric oxide (NO) and oleic acid (18:1) are well-known regulators of disease physiologies in diverse organism. We show that NO production in plants is regulated via 18:1. Reduction in 18:1 levels, via a genetic mutation in the 18:1-synthesizing gene SUPPRESSOR OF S...

  12. Protein biosynthesis with conformationally restricted amino acids

    SciTech Connect

    Mendel, D. Lawrence Berkeley Lab., CA ); Ellman, J.; Schultz, P.G. )

    1993-05-19

    The incorporation of conformationally constrained amino acids into peptides is a powerful approach for generating structurally defined peptides as conformational probes and bioactive agents. The ability to site-specifically introduce constrained amino acids into large polypeptide chains would provide a similar opportunity to probe the flexibility, conformation, folding and stability of proteins. To this end, we have examined the competence of the Escherichia coli protein biosynthetic machinery to incorporate a number of these unnatural amino acids into the 164 residue protein T4 lysozyme (T4L). Results clearly demonstrate that the protein biosynthetic machinery can accommodate a wide variety of conformationally constrained amino acids. The expansion of structural motifs that can be biosynthetically incorporated into proteins to include a large number of conformationally constrained amino acids significantly increases the power of mutagenesis methods as probes of protein structure and function and provides additional insights into the steric requirements of the translational machinery. 13 refs., 2 figs.

  13. Distinguishing proteins from arbitrary amino acid sequences.

    PubMed

    Yau, Stephen S-T; Mao, Wei-Guang; Benson, Max; He, Rong Lucy

    2015-01-01

    What kinds of amino acid sequences could possibly be protein sequences? From all existing databases that we can find, known proteins are only a small fraction of all possible combinations of amino acids. Beginning with Sanger's first detailed determination of a protein sequence in 1952, previous studies have focused on describing the structure of existing protein sequences in order to construct the protein universe. No one, however, has developed a criteria for determining whether an arbitrary amino acid sequence can be a protein. Here we show that when the collection of arbitrary amino acid sequences is viewed in an appropriate geometric context, the protein sequences cluster together. This leads to a new computational test, described here, that has proved to be remarkably accurate at determining whether an arbitrary amino acid sequence can be a protein. Even more, if the results of this test indicate that the sequence can be a protein, and it is indeed a protein sequence, then its identity as a protein sequence is uniquely defined. We anticipate our computational test will be useful for those who are attempting to complete the job of discovering all proteins, or constructing the protein universe. PMID:25609314

  14. Liver Fatty Acid Binding Protein and Obesity

    PubMed Central

    Atshaves, B.P.; Martin, G.G.; Hostetler, H.A.; McIntosh, A.L.; Kier, A.B.; Schroeder, F.

    2010-01-01

    While low levels of unesterified long chain fatty acids (LCFAs) are normal metabolic intermediates of dietary and endogenous fat, LCFAs are also potent regulators of key receptors/enzymes, and at high levels become toxic detergents within the cell. Elevated levels of LCFAs are associated with diabetes, obesity, and metabolic syndrome. Consequently, mammals evolved fatty acid binding proteins (FABPs) that bind/sequester these potentially toxic free fatty acids in the cytosol and present them for rapid removal in oxidative (mitochondria, peroxisomes) or storage (endoplasmic reticulum, lipid droplets) organelles. Mammals have a large (15 member) family of FABPs with multiple members occurring within a single cell type. The first described FABP, liver-FABP (L-FABP, or FABP1), is expressed in very high levels (2-5% of cytosolic protein) in liver as well as intestine and kidney. Since L-FABP facilitates uptake and metabolism of LCFAs in vitro and in cultured cells, it was expected that abnormal function or loss of L-FABP would reduce hepatic LCFA uptake/oxidation and thereby increase LCFAs available for oxidation in muscle and/or storage in adipose. This prediction was confirmed in vitro with isolated liver slices and cultured primary hepatocytes from L-FABP gene-ablated mice. Despite unaltered food consumption when fed a control diet ad libitum, the L-FABP null mice exhibited age- and sex-dependent weight gain and increased fat tissue mass. The obese phenotype was exacerbated in L-FABP null mice pair-fed a high fat diet. Taken together with other findings, these data suggest that L-FABP could have an important role in preventing age- or diet-induced obesity. PMID:20537520

  15. Protein, amino acids and the control of food intake.

    PubMed

    Tome, Daniel

    2004-08-01

    The influence of protein and amino acid on the control of food intake and the specific control of protein and amino acid intakes remains incompletely understood. The most commonly accepted conclusions are: (1) the existence of an aversive response to diets deficient in or devoid of protein or deficient in at least one essential amino acid; (2) the existence of a mechanism that enables attainment of the minimum requirement for N and essential amino acids by increasing intake of a low-protein diet; (3) a decrease in the intake of a high-protein diet is associated with different processes, including the high satiating effect of protein. Ingested proteins are believed to generate pre- and post-absorptive signals that contribute to the control of gastric kinetics, pancreatic secretion and food intake. At the brain level, two major afferent pathways are involved in protein and amino acid monitoring: the indirect neuro-mediated (mainly vagus-mediated) pathway and the direct blood pathway. The neuro-mediated pathway transfers pre-absorptive and visceral information. This information is for the main part transferred through the vagus nerve that innervates part of the oro-sensory zone: the stomach, the duodenum and the liver. Other information is directly monitored in the blood. It is likely that the system responds precisely when protein and essential amino acid intake is inadequate, but in contrast allows a large range of adaptive capacities through amino acid degradation and substrate interconversion.

  16. Effect of aspartame plus monosodium L-glutamate ingestion on plasma and erythrocyte amino acid levels in normal adult subjects fed a high protein meal.

    PubMed

    Stegink, L D; Filer, L J; Baker, G L

    1982-12-01

    It has been suggested that aspartame addition to meals already containing large amounts of monosodium L-glutamate would result in an early rapid rise in plasma glutamate and/or aspartate concentrations and increase the potential for dicarboxylic amino acid-induced toxicity. Six normal adult subjects were fed hamburger and milk shake meals providing protein at 1 g/kg body weight in a randomized cross-over design. One meal had no additions while the other contained added monosodium L-glutamate and aspartame (each present at 34 mg/kg body weight). The addition of aspartame plus glutamate had little effect on either plasma or erythrocyte concentrations of glutamate or aspartate beyond those arising from the meal itself. Plasma phenylalanine concentrations were significantly higher (p less than 0.05, paired t test) after ingestion of meals containing aspartame plus glutamate reflecting the increased phenylalanine load.

  17. A new acidic protein in porcine brain.

    PubMed

    Ishioka, N; Isobe, T; Okuyama, T; Numata, Y; Wada, H

    1980-10-21

    An extremely acidic protein has been isolated in a purified form from porcine rain extract, by (NH4)2SO4 fractionation followed by column chromatography on DEAE-Sephadex A-50 and on Sephadex G-75. The purified protein was tentatively named as glutamic acid-rich protein because it was characterized by its remarkably high content of glutamic acid which accounted for 49% of the total amino acid composition. The protein appeared to be a single polypeptide chain with a molecular weight of 56 000-58 000, and had an isoelectric point of 4.6. The N-terminal amino acid sequence was Asp-Glu-Pro-Pro-Ser-Glu-Gly. The immunochemical analysis using rabbit antiserum prepared to the porcine protein has suggested that it is present in the brain of human, cow, cat, dog and goat as well as in various goat organs including liver, kidney, heart, small intestine and spleen.

  18. [Abscisic acid level, activities of proteinases and trypsin inhibitory proteins in the germinating seeds of common beans under conditions of water stress].

    PubMed

    Domash, V I; Protsko, R F; Vasiuk, V A; Shumikhin, S V; Ermolitskaia, L V; Sharpio, T P

    2006-01-01

    Specific features of changes in the contents of free and bound abscisic acid (ABA) and activities of neutral and alkaline proteinases and trypsin inhibitory proteins were determined in the embryonic axis and cotyledons of the common bean (Phaseolus vulgaris L.) after drying. The changes in ABA content, observed following the loss of 5% seed weight, were regarded as an adaptive reaction to stress, whereas the corresponding changes after the loss of 10% seed weight, as a result of pathological disturbance of ABA metabolism. Both drying modes had a negative effect on the state of the proteinase-inhibitory system, as was apparent from the disruption of the regular inverse correlation between the activities of proteinases and serine proteinase inhibitory proteins. Comparison of the dynamics of these characteristics with the buildup of water stress demonstrated an inverse correlation between the content of free ABA and the activity of the proteinases studied. This suggests a potential inhibitory effect of this hormone on the function of the hydrolases in question in the germinating seed.

  19. BIOPOLYMERS FROM POLYLACTIC ACID AND MILK PROTEINS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polylactic acid (PLA) is a commercially available biodegradable polymer derived from lactic acid and is used in many nonfood products as an alternative to petrochemical-derived polymers. However, its physical properties limit its use in many applications. Using dairy proteins to substitute for por...

  20. The effects of orbital spaceflight on bone histomorphometry and messenger ribonucleic acid levels for bone matrix proteins and skeletal signaling peptides in ovariectomized growing rats

    NASA Technical Reports Server (NTRS)

    Cavolina, J. M.; Evans, G. L.; Harris, S. A.; Zhang, M.; Westerlind, K. C.; Turner, R. T.

    1997-01-01

    A 14-day orbital spaceflight was performed using ovariectomized Fisher 344 rats to determine the combined effects of estrogen deficiency and near weightlessness on tibia radial bone growth and cancellous bone turnover. Twelve ovariectomized rats with established cancellous osteopenia were flown aboard the space shuttle Columbia (STS-62). Thirty ovariectomized rats were housed on earth as ground controls: 12 in animal enclosure modules, 12 in vivarium cages, and 6 killed the day of launch for baseline measurements. An additional 18 ovary-intact rats were housed in vivarium cages as ground controls: 8 rats were killed as baseline controls and the remaining 10 rats were killed 14 days later. Ovariectomy increased periosteal bone formation at the tibia-fibula synostosis; cancellous bone resorption and formation in the secondary spongiosa of the proximal tibial metaphysis; and messenger RNA (mRNA) levels for the prepro-alpha2(1) subunit of type 1 collagen, osteocalcin, transforming growth factor-beta, and insulin-like growth factor I in the contralateral proximal tibial metaphysis and for the collagen subunit in periosteum pooled from tibiae and femora and decreased cancellous bone area. Compared to ovariectomized weight-bearing rats, the flight group experienced decreases in periosteal bone formation, collagen subunit mRNA levels, and cancellous bone area. The flight rats had a small decrease in the cancellous mineral apposition rate, but no change in the calculated bone formation rate. Also, spaceflight had no effect on cancellous osteoblast and osteoclast perimeters or on mRNA levels for bone matrix proteins and signaling peptides. On the other hand, spaceflight resulted in an increase in bone resorption, as ascertained from the diminished retention of a preflight fluorochrome label. This latter finding suggests that osteoclast activity was increased. In a follow-up ground-based experiment, unilateral sciatic neurotomy of ovariectomized rats resulted in cancellous

  1. Probing protein stability with unnatural amino acids

    SciTech Connect

    Mendel, D.; Ellman, J.A.; Zhiyuh Chang; Veenstra, D.L.; Kollman, P.A.; Schultz, P.G. )

    1992-06-26

    Unnatural amino acid mutagenesis, in combination with molecular modeling and simulation techniques, was used to probe the effect of side chain structure on protein stability. Specific replacements at position 133 in T4 lysozyme included (1) leucine (wt), norvaline, ethylglycine, and alanine to measure the cost of stepwise removal of methyl groups from the hydrophobic core, (2) norvaline and O-methyl serine to evaluate the effects of side chain solvation, and (3) leucine, S,S-2-amino-4-methylhexanoic acid, and S-2-amino-3-cyclopentylpropanoic acid to measure the influence of packing density and side chain conformational entropy on protein stability. All of these factors (hydrophobicity, packing, conformational entropy, and cavity formation) significantly influence protein stability and must be considered when analyzing any structural change to proteins.

  2. Detection of non-protein amino acids in the presence of protein amino acids. II.

    NASA Technical Reports Server (NTRS)

    Shapshak, P.; Okaji, M.

    1972-01-01

    Studies conducted with the JEOL 5AH amino acid analyzer are described. This instrument makes possible the programming of the chromatographic process. Data are presented showing the separations of seventeen non-protein amino acids in the presence of eighteen protein amino acids. It is pointed out that distinct separations could be obtained in the case of a number of chemically similar compounds, such as ornithine and lysine, N-amidino alanine and arginine, and iminodiacetic acid and S-carboxymethyl cysteine and aspartic acid.

  3. Nucleic acids, proteins, and chirality

    NASA Technical Reports Server (NTRS)

    Usher, D. A.; Profy, A. T.; Walstrum, S. A.; Needels, M. C.; Bulack, S. C.; Lo, K. M.

    1984-01-01

    The present investigation is concerned with experimental results related, in one case, to the chirality of nucleotides, and, in another case, to the possibility of a link between the chirality of nucleic acids, and that of peptides. It has been found that aminoacylation of the 'internal' hydroxyl group of a dinucleoside monophosphate can occur stereoselectively. However, this reaction has not yet been made a part of a working peptide synthesis scheme. The formation and cleavage of oligonucleotides is considered. In the event of the formation of a helical complex between the oligonucleotide and the polymer, 1-prime,5-prime-bonds in the oligomer are found to become more resistant towards cleavage. The conditions required for peptide bond formation are examined, taking into account the known structures of RNA and possible mechanisms for prebiotic peptide bond formation. The possibility is considered that the 2-prime,5-prime-internucleotide linkage could have played an important part in the early days of biological peptide synthesis.

  4. The calcium-dependent protein kinase CPK28 regulates development by inducing growth phase-specific, spatially restricted alterations in jasmonic acid levels independent of defense responses in Arabidopsis.

    PubMed

    Matschi, Susanne; Hake, Katharina; Herde, Marco; Hause, Bettina; Romeis, Tina

    2015-03-01

    Phytohormones play an important role in development and stress adaptations in plants, and several interacting hormonal pathways have been suggested to accomplish fine-tuning of stress responses at the expense of growth. This work describes the role played by the CALCIUM-DEPENDENT PROTEIN KINASE CPK28 in balancing phytohormone-mediated development in Arabidopsis thaliana, specifically during generative growth. cpk28 mutants exhibit growth reduction solely as adult plants, coinciding with altered balance of the phytohormones jasmonic acid (JA) and gibberellic acid (GA). JA-dependent gene expression and the levels of several JA metabolites were elevated in a growth phase-dependent manner in cpk28, and accumulation of JA metabolites was confined locally to the central rosette tissue. No elevated resistance toward herbivores or necrotrophic pathogens was detected for cpk28 plants, either on the whole-plant level or specifically within the tissue displaying elevated JA levels. Abolishment of JA biosynthesis or JA signaling led to a full reversion of the cpk28 growth phenotype, while modification of GA signaling did not. Our data identify CPK28 as a growth phase-dependent key negative regulator of distinct processes: While in seedlings, CPK28 regulates reactive oxygen species-mediated defense signaling; in adult plants, CPK28 confers developmental processes by the tissue-specific balance of JA and GA without affecting JA-mediated defense responses.

  5. The Calcium-Dependent Protein Kinase CPK28 Regulates Development by Inducing Growth Phase-Specific, Spatially Restricted Alterations in Jasmonic Acid Levels Independent of Defense Responses in Arabidopsis[OPEN

    PubMed Central

    Matschi, Susanne; Hake, Katharina; Herde, Marco; Hause, Bettina; Romeis, Tina

    2015-01-01

    Phytohormones play an important role in development and stress adaptations in plants, and several interacting hormonal pathways have been suggested to accomplish fine-tuning of stress responses at the expense of growth. This work describes the role played by the CALCIUM-DEPENDENT PROTEIN KINASE CPK28 in balancing phytohormone-mediated development in Arabidopsis thaliana, specifically during generative growth. cpk28 mutants exhibit growth reduction solely as adult plants, coinciding with altered balance of the phytohormones jasmonic acid (JA) and gibberellic acid (GA). JA-dependent gene expression and the levels of several JA metabolites were elevated in a growth phase-dependent manner in cpk28, and accumulation of JA metabolites was confined locally to the central rosette tissue. No elevated resistance toward herbivores or necrotrophic pathogens was detected for cpk28 plants, either on the whole-plant level or specifically within the tissue displaying elevated JA levels. Abolishment of JA biosynthesis or JA signaling led to a full reversion of the cpk28 growth phenotype, while modification of GA signaling did not. Our data identify CPK28 as a growth phase-dependent key negative regulator of distinct processes: While in seedlings, CPK28 regulates reactive oxygen species-mediated defense signaling; in adult plants, CPK28 confers developmental processes by the tissue-specific balance of JA and GA without affecting JA-mediated defense responses. PMID:25736059

  6. Association of Renal Manifestations with Serum Uric Acid in Korean Adults with Normal Uric Acid Levels

    PubMed Central

    Jung, Dong-Hyuk; Lee, Yong-Jae; Lee, Hye-Ree; Lee, Jung-Hyun

    2010-01-01

    Several studies have reported that hyperuricemia is associated with the development of hypertension and cardiovascular disease. Increasing evidences also suggest that hyperuricemia may have a pathogenic role in the progression of renal disease. Paradoxically, uric acid is also widely accepted to have antioxidant activity in experimental studies. We aimed to investigate the association between glomerular filtration rate (GFR) and uric acid in healthy individuals with a normal serum level of uric acid. We examined renal function determined by GFR and uric acid in 3,376 subjects (1,896 men; 1,480 women; aged 20-80 yr) who underwent medical examinations at Gangnam Severance Hospital from November 2006 to June 2007. Determinants for renal function and uric acid levels were also investigated. In both men and women, GFR was negatively correlated with systolic and diastolic blood pressures, fasting plasma glucose, total cholesterol, uric acid, log transformed C reactive protein, and log transformed triglycerides. In multivariate regression analysis, total uric acid was found to be an independent factor associated with estimated GFR in both men and women. This result suggests that uric acid appears to contribute to renal impairment in subjects with normal serum level of uric acid. PMID:21165292

  7. Alimentary proteins, amino acids and cholesterolemia.

    PubMed

    Blachier, François; Lancha, Antonio H; Boutry, Claire; Tomé, Daniel

    2010-01-01

    Numerous data from both epidemiological and experimental origins indicate that some alimentary proteins and amino acids in supplements can modify the blood LDL cholesterol, HDL cholesterol and total cholesterol. After an initial approval of the health claim for soy protein consumption for the prevention of coronary heart disease, more recently it has been concluded from an overall analysis of literature that isolated soy protein with isoflavones only slightly decrease LDL and total cholesterol. Other plant extracts and also some proteins from animal origin have been reported to exert a lowering effect on blood cholesterol when compared with a reference protein (often casein). The underlying mechanisms are still little understood. Individual amino acids and mixture of amino acids have also been tested (mostly in animal studies) for their effects on cholesterol parameters and on cholesterol metabolism. Methionine, lysine, cystine, leucine, aspartate and glutamate have been tested individually and in combination in different models of either normo or hypercholesterolemic animals and found to be able to modify blood cholesterol and/or LDL cholesterol and/or HDL cholesterol. It is however not known if these results are relevant to human nutrition.

  8. Role of fatty acid binding protein on hepatic palmitate uptake.

    PubMed

    Burczynski, F J; Zhang, M N; Pavletic, P; Wang, G Q

    1997-12-01

    Expression of hepatic fatty acid binding protein (FABP) mRNA is regulated by growth hormone. In the absence of growth hormone, there is a 60% reduction in FABP mRNA levels (S.A. Berry, J.-B Yoon, U. List, and S. Seelig. J. Am. Coll. Nutr. 12:638-642. 1995). Previous work in our laboratory focused on the role of extracellular binding proteins in the hepatic uptake of long chain fatty acids. In the present study we were interested to determine the role of FABP in the transmembrane flux of long chain fatty acids. Using hepatocyte monolayers from control (n = 9) and hypophysectomized (n = 6) rats, we investigated the uptake of [3H]palmitate in the presence and absence of albumin. In the absence of albumin, total hepatocyte [3H]palmitate clearance rates from control (17.2 +/- 1.5 microL.mg-1 protein.s-1; mean +/- SEM; n = 9) and hypophysectomized (15.5 +/- 2.1 microL.mg-1 protein.s-1; n = 6) animals were similar (p > 0.05). In the presence of 2 microM albumin the total [3H]palmitate clearance rate from control hepatocytes (1.63 +/- 0.11 microL.mg-1 protein.s-1; n = 9) was significantly larger (40%) than from hepatocytes obtained from hypophysectomized (0.97 +/- 0.15 microL.mg-1 protein.s-1; n = 6; p < 0.01) animals. SDS-PAGE electrophoresis revealed that plasma membrane FABP levels from control and hypophysectomized animals were similar. However, there was a 49% decrease in the cytosolic FABP levels of hepatocytes isolated from hypophysectomized as compared with control animals. The decreased cytosolic FABB levels paralleled the decrease in palmitate uptake. We conclude that in the absence of extracellular binding proteins the rate-limiting step in the overall uptake of long chain fatty acids is diffusion to the cell surface. However, in the presence of albumin, the rate of palmitate uptake is determined primarily by cytosolic FABP levels.

  9. (-)-Hydroxycitric Acid Nourishes Protein Synthesis via Altering Metabolic Directions of Amino Acids in Male Rats.

    PubMed

    Han, Ningning; Li, Longlong; Peng, Mengling; Ma, Haitian

    2016-08-01

    (-)-Hydroxycitric acid (HCA), a major active ingredient of Garcinia Cambogia extracts, had shown to suppress body weight gain and fat accumulation in animals and humans. While, the underlying mechanism of (-)-HCA has not fully understood. Thus, this study was aimed to investigate the effects of long-term supplement with (-)-HCA on body weight gain and variances of amino acid content in rats. Results showed that (-)-HCA treatment reduced body weight gain and increased feed conversion ratio in rats. The content of hepatic glycogen, muscle glycogen, and serum T4 , T3 , insulin, and Leptin were increased in (-)-HCA treatment groups. Protein content in liver and muscle were significantly increased in (-)-HCA treatment groups. Amino acid profile analysis indicated that most of amino acid contents in serum and liver, especially aromatic amino acid and branched amino acid, were higher in (-)-HCA treatment groups. However, most of the amino acid contents in muscle, especially aromatic amino acid and branched amino acid, were reduced in (-)-HCA treatment groups. These results indicated that (-)-HCA treatment could reduce body weight gain through promoting energy expenditure via regulation of thyroid hormone levels. In addition, (-)-HCA treatment could promote protein synthesis by altering the metabolic directions of amino acids. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27145492

  10. Protein and Amino Acid Requirements during Pregnancy.

    PubMed

    Elango, Rajavel; Ball, Ronald O

    2016-07-01

    Protein forms an essential component of a healthy diet in humans to support both growth and maintenance. During pregnancy, an exceptional stage of life defined by rapid growth and development, adequate dietary protein is crucial to ensure a healthy outcome. Protein deposition in maternal and fetal tissues increases throughout pregnancy, with most occurring during the third trimester. Dietary protein intake recommendations are based on factorial estimates because the traditional method of determining protein requirements, nitrogen balance, is invasive and undesirable during pregnancy. The current Estimated Average Requirement and RDA recommendations of 0.88 and 1.1 g · kg(-1) · d(-1), respectively, are for all stages of pregnancy. The single recommendation does not take into account the changing needs during different stages of pregnancy. Recently, with the use of the minimally invasive indicator amino acid oxidation method, we defined the requirements to be, on average, 1.2 and 1.52 g · kg(-1) · d(-1) during early (∼16 wk) and late (∼36 wk) stages of pregnancy, respectively. Although the requirements are substantially higher than current recommendations, our values are ∼14-18% of total energy and fit within the Acceptable Macronutrient Distribution Range. Using swine as an animal model we showed that the requirements for several indispensable amino acids increase dramatically during late gestation compared with early gestation. Additional studies should be conducted during pregnancy to confirm the newly determined protein requirements and to determine the indispensable amino acid requirements during pregnancy in humans. PMID:27422521

  11. Acidic Shell Proteins of the Mediterranean Fan Mussel Pinna nobilis.

    PubMed

    Marin, Frédéric; Narayanappa, Prabakaran; Motreuil, Sébastien

    2011-01-01

    In molluscs, the shell secretion process is controlled by a set of extracellular macromolecules collectively called the shell matrix. The shell matrix, which is produced by the mantle epithelial cells during mineralization, is predominantly composed of proteins, glycoproteins, acidic polysaccharides, and chitin that precisely regulate the deposition of calcium carbonate outside the mantle cells. In the present paper, we focus on the shell of Pinna nobilis, the giant Mediterranean fan mussel, usually considered as a model for studying molluscan biomineralization processes. P. nobilis exhibits indeed a nacro-prismatic shell, the outer layer of which is constituted of the so-called "regular simple calcitic prisms," according to Carter and Clark (1985). We review here the microstructural characteristics of the prisms and nacre and the biochemical properties of their associated matrices. In particular, the calcitic prisms of P. nobilis are characterized by a cortege of unusually acidic intraprismatic proteins, while the ones of the nacreous layer seem less acidic. A brief description of the molecular characterization of three acidic proteins, caspartin, calprismin and mucoperlin, is given. In particular, we show that extremely acidic intracrystalline proteins such as caspartin interact with calcium carbonate at different scales, from micrometric to crystal lattice levels.

  12. Heavy metals influence on ascorbic acid level

    NASA Astrophysics Data System (ADS)

    Kamaldinov, E. V.; Patrashkov, S. A.; Batenyeva, E. V.; Korotkevich, O. S.

    2003-05-01

    It is well known that heavy metals (HM) are extremely dangerous pollutants influencing to metabolism in animals' organisms. The vitamin C is one of the most important metabolites taking part in many biochemical processes. We studied the influence of main essential HM-Zn and Cu as well as the based supertoxical elements - Cd and Pd on ascorbic acid level in serum. The studies were carried out in Tulinskoe farm of Novosibirsk region. The objects of investigations were piglets (2 month after weaning) and 6-month pigs of Early Ripe Meat breed. The levels of HM in bristle were found by stripping voltammetric analysis using the TA-2 analyzer. Vitamin C content was determined by I.P. Kondrakhin (1985) method using 2,2-dipyridyl. The significant negative correlations between Pb, Cd content and vitamin C (-0.46 ± 0.18, -0.47 ± 0.19) in 6-month pigs were determined. The tendencies of negative correlation between all HM levels in hair and ascorbic acid level in plasma of piglets were revealed. Thus, the obtained correlations let us to suppose that all studied HM influence on 1-gulono-gamma-lactone oxidase and other vitamin C metabolism enzymes activity.

  13. Free amino acids in crocodilians fed proteins of different biological value.

    PubMed

    Herbert, J D; Coulson, R A

    1975-05-01

    Changes in plasma levels of amino acids derived from fed protein were determined by feeding crocodilians (Caiman crocodilus crocodilus and Alligator mississipiensis) 7.5 g protein/kg body weight and by monitoring the plasma free amino acids for several days. Zein and several other vegetable proteins produced no rise in plasma amino acids and were excreted intact in the feces. Casein and fish muscle were rapidly digested but produced little rise in plasma amino acids, and the increases showed no relationship to the composition of the protein fed. Gelatin feeding led to large increases in plasma amino acids that persisted for more than a week, and the resulting pattern was nearly identical to the composition of gelatin with the exception of aspartic and glutamic acids, and several animals died. Equivalent quantities of fish muscle protein were assimilated without difficulty by the crocodilians. Endogenous protein secreted into the gut apparently contributed little to the amino acid mixture absorbed.

  14. Amino Acid Recycling in Relation to Protein Turnover 1

    PubMed Central

    Davies, David D.; Humphrey, Thomas J.

    1978-01-01

    Methods of measuring amino acid recycling in Lemna minor are described. The extent to which the recycling of individual amino acids may underestimate protein turnover has been measured for a number of amino acids. The methods have been used to study the relationship between protein turnover and amino acid recycling during nitrogen starvation. It is concluded that following the removal of nitrate from the environment, protein turnover is enhanced, the partitioning of amino acids between protein synthesis and amino acid metabolism is relatively constant, but the total amount of amino acids recycling is increased. PMID:16660236

  15. Transcriptional regulation of muscle fatty acid-binding protein.

    PubMed Central

    Carey, J O; Neufer, P D; Farrar, R P; Veerkamp, J H; Dohm, G L

    1994-01-01

    Heart fatty acid-binding protein (H-FABP) is present in a wide variety of tissues but is found in the highest concentration in cardiac and red skeletal muscle. It has been proposed that the expression of H-FABP correlates directly with the fatty acid-oxidative capacity of the tissue. In the present study, the expression of H-FABP was measured in red and white skeletal muscle under two conditions in which fatty acid utilization is known to be increased: streptozotocin-induced diabetes and fasting. Protein concentration, mRNA concentration and transcription rate were measured under both conditions. The level of both protein and mRNA increased approximately 2-fold under each condition. The transcription rate was higher in red skeletal muscle than in white muscle, was increased 2-fold during fasting, but was unchanged by streptozotocin-induced diabetes. In addition to supporting the hypothesis that H-FABP is induced during conditions of increased fatty acid utilization, these findings demonstrate that the regulation of H-FABP expression may or may not be at the level of transcription depending on the stimulus. Images Figure 2 Figure 3 PMID:8141774

  16. Protein and Amino Acid Profiles of Different Whey Protein Supplements.

    PubMed

    Almeida, Cristine C; Alvares, Thiago S; Costa, Marion P; Conte-Junior, Carlos A

    2016-01-01

    Whey protein (WP) supplements have received increasing attention by consumers due to the high nutritional value of the proteins and amino acids they provide. However, some WP supplements may not contain the disclosed amounts of the ingredients listed on the label, compromising the nutritional quality and the effectiveness of these supplements. The aim of this study was to evaluate and compare the contents of total protein (TP), α-lactalbumin (α-LA), β-lactoglobulin (β-LG), free essential amino acids (free EAA), and free branched-chain amino acids (free BCAA), amongst different WP supplements produced by U.S. and Brazilian companies. Twenty commercial brands of WP supplements were selected, ten manufactured in U.S. (WP-USA) and ten in Brazil (WP-BRA). The TP was analyzed using the Kjeldahl method, while α-LA, β-LG, free EAA, and free BCAA were analyzed using HPLC system. There were higher (p < 0.05) concentrations of TP, α-LA, β-LG, and free BCAA in WP-USA supplements, as compared to the WP-BRA supplements; however, there was no difference (p > 0.05) in the content of free EAA between WP-USA and WP-BRA. Amongst the 20 brands evaluated, four WP-USA and seven WP-BRA had lower (p < 0.05) values of TP than those specified on the label. In conclusion, the WP-USA supplements exhibited better nutritional quality, evaluated by TP, α-LA, β-LG, and free BCAA when compared to WP-BRA.

  17. Abscisic Acid Levels and Seed Dormancy

    PubMed Central

    Sondheimer, E.; Tzou, D. S.; Galson, Eva C.

    1968-01-01

    Dormant seeds from Fraxinus species require cold-temperature after-ripening prior to germination. Earlier, we found that abscisic acid (ABA) will inhibit germination of excised nondormant embryos and that this can be reversed with a combination of gibberellic acid and kinetin. Using Milborrow's quantitative “racemate dilution” method the ABA concentration in 3 types of Fraxinus seed and pericarp were determined. While ABA was present in all tissues, the highest concentration was found in the seed and pericarp of dormant F. americana. During the chilling treatment of F. americana the ABA levels decreased 37% in the pericarp and 68% in the seed. The ABA concentration of the seed of the nondormant species, F. ornus, is as low as that found in F. americana seeds after cold treatment. Experiments with exogenously added ABA solutions indicate that it is unlikely that the ABA in the pericarp functions in the regulation of seed dormancy. However, the ABA in the seed does seem to have a regulatory role in germination. Images PMID:16656935

  18. Residual protein levels on reprocessed dental instruments.

    PubMed

    Smith, A; Letters, S; Lange, A; Perrett, D; McHugh, S; Bagg, J

    2005-11-01

    Reduction of the initial bioburden on instruments, prior to sterilization, is believed to reduce transmission risks of iatrogenic Creutzfeldt-Jakob disease. Endodontic files are used in the preparation of root canals and are likely to have close contact and become contaminated with neural material from branches of the maxillary and mandibular cranial nerves. This study examined methods used by 22 dental practices to clean endodontic files, and scored visible debris and residual protein levels adhering to 220 dental endodontic files that had been used, cleaned, autoclaved and were deemed ready for re-use. Visible debris was scored after examination under a dissecting light microscope. Residual protein was quantified using a fluorescent assay based on reaction of proteins with o-phthaldialdehyde/N-acetyl cysteine. There was wide variation in the methods used by practices to clean endodontic files. The cleaning process varied from a wipe with an alcohol-impregnated cloth to hand scrubbing and/or use of an ultrasonic bath. Surface debris was visually detected on 98% of files. Residual protein was detected on all the files examined (median amount: 5.4 microg; range: 0.5-63.2 microg). These results demonstrate that the cleaning of some instruments reprocessed routinely in primary care is incomplete, and such instruments cannot be excluded as a potential source of cross-infection.

  19. Classification of G-protein coupled receptors at four levels.

    PubMed

    Gao, Qing-Bin; Wang, Zheng-Zhi

    2006-11-01

    G-protein coupled receptors (GPCRs) are transmembrane proteins which via G-proteins initiate some of the important signaling pathways in a cell and are involved in various physiological processes. Thus, computational prediction and classification of GPCRs can supply significant information for the development of novel drugs in pharmaceutical industry. In this paper, a nearest neighbor method has been introduced to discriminate GPCRs from non-GPCRs and subsequently classify GPCRs at four levels on the basis of amino acid composition and dipeptide composition of proteins. Its performance is evaluated on a non-redundant dataset consisted of 1406 GPCRs for six families and 1406 globular proteins using the jackknife test. The present method based on amino acid composition achieved an overall accuracy of 96.4% and Matthew's correlation coefficient (MCC) of 0.930 for correctly picking out the GPCRs from globular proteins. The overall accuracy and MCC were further enhanced to 99.8% and 0.996 by dipeptide composition-based method. On the other hand, the present method has successfully classified 1406 GPCRs into six families with an overall accuracy of 89.6 and 98.8% using amino acid composition and dipeptide composition, respectively. For the subfamily prediction of 1181 GPCRs of rhodopsin-like family, the present method achieved an overall accuracy of 76.7 and 94.5% based on the amino acid composition and dipeptide composition, respectively. Finally, GPCRs belonging to the amine subfamily and olfactory subfamily of rhodopsin-like family were further analyzed at the type level. The overall accuracy of dipeptide composition-based method for the classification of amine type and olfactory type of GPCRs reached 94.5 and 86.9%, respectively, while the overall accuracy of amino acid composition-based method was very low for both subfamilies. In comparison with existing methods in the literature, the present method also displayed great competitiveness. These results demonstrate

  20. Impact of antinutritional factors in food proteins on the digestibility of protein and the bioavailability of amino acids and on protein quality.

    PubMed

    Sarwar Gilani, G; Wu Xiao, Chao; Cockell, Kevin A

    2012-08-01

    Dietary antinutritional factors have been reported to adversely affect the digestibility of protein, bioavailability of amino acids and protein quality of foods. Published data on these negative effects of major dietary antinutritional factors are summarized in this manuscript. Digestibility and the quality of mixed diets in developing countries are considerably lower than of those in developed regions. For example, the digestibility of protein in traditional diets from developing countries such as India, Guatemala and Brazil is considerably lower compared to that of protein in typical North American diets (54-78 versus 88-94 %). Poor digestibility of protein in the diets of developing countries, which are based on less refined cereals and grain legumes as major sources of protein, is due to the presence of less digestible protein fractions, high levels of insoluble fibre, and/or high concentrations of antinutritional factors present endogenously or formed during processing. Examples of naturally occurring antinutritional factors include glucosinolates in mustard and canola protein products, trypsin inhibitors and haemagglutinins in legumes, tannins in legumes and cereals, gossypol in cottonseed protein products, and uricogenic nucleobases in yeast protein products. Heat/alkaline treatments of protein products may yield Maillard reaction compounds, oxidized forms of sulphur amino acids, D-amino acids and lysinoalanine (LAL, an unnatural nephrotoxic amino acid derivative). Among common food and feed protein products, soyabeans are the most concentrated source of trypsin inhibitors. The presence of high levels of dietary trypsin inhibitors from soyabeans, kidney beans or other grain legumes have been reported to cause substantial reductions in protein and amino acid digestibility (up to 50 %) and protein quality (up to 100 %) in rats and/or pigs. Similarly, the presence of high levels of tannins in sorghum and other cereals, fababean and other grain legumes can cause

  1. Nucleic acids encoding human trithorax protein

    DOEpatents

    Evans, Glen A.; Djabali, Malek; Selleri, Licia; Parry, Pauline

    2001-01-01

    In accordance with the present invention, there is provided an isolated peptide having the characteristics of human trithorax protein (as well as DNA encoding same, antisense DNA derived therefrom and antagonists therefor). The invention peptide is characterized by having a DNA binding domain comprising multiple zinc fingers and at least 40% amino acid identity with respect to the DNA binding domain of Drosophila trithorax protein and at least 70% conserved sequence with respect to the DNA binding domain of Drosophila trithorax protein, and wherein said peptide is encoded by a gene located at chromosome 11 of the human genome at q23. Also provided are methods for the treatment of subject(s) suffering from immunodeficiency, developmental abnormality, inherited disease, or cancer by administering to said subject a therapeutically effective amount of one of the above-described agents (i.e., peptide, antagonist therefor, DNA encoding said peptide or antisense DNA derived therefrom). Also provided is a method for the diagnosis, in a subject, of immunodeficiency, developmental abnormality, inherited disease, or cancer associated with disruption of chromosome 11 at q23.

  2. Response of protein and urea kinetics in burn patients to different levels of protein intake.

    PubMed Central

    Wolfe, R R; Goodenough, R D; Burke, J F; Wolfe, M H

    1983-01-01

    The effects of two levels of protein intake on protein metabolism in six severely burned adult patients were studied (means of 70% BSA burned). A crossover experimental design enabled the authors to study each patient at the end of two three-day dietary regimens. All diets were isocaloric and provided approximately 25% more calories than the measured energy expenditure (means = 40.8 Kcal/kg X day). In one regimen, each patient received 2.2 g protein/kg X day, while during the other treatment period they received 1.4 g protein/kg X day. The patients were studied in the fed state and after 10 to 12 hours of fasting. Leucine kinetics were determined by means of the primed-constant infusion of [1--13C]--leucine. The authors were able to distinguish the oxidation of plasma leucine from the oxidation of leucine derived from intracellular protein at the site of the deamination of leucine (predominantly muscle) by simultaneously determining both leucine and alpha-ketoisocaproic acid enrichment. Also, rates of whole-body protein synthesis and catabolism were calculated from the leucine flux and oxidation data. Net protein synthesis was also calculated by means of another stable-isotope technique involving the infusion of [15N2]--urea. Finally, a third means of estimating net protein catabolism based on urinary N-excretion data was used at the same time that the isotopic studies were performed. The 13C leucine-data and the N-excretion data indicated that a balance between protein synthesis and catabolism could be achieved with a protein intake of 1.4 protein/kg X day. When protein intake was increased to 2.2 g protein/kg X day, neither isotopic method indicated a further beneficial effect on net protein synthesis, although the absolute rates of protein synthesis and catabolism were stimulated. The N-excretion data, on the other hand, indicated a significant improvement in net protein synthesis with higher protein intake. Regardless of the level of protein intake, the

  3. ORMDL proteins regulate ceramide levels during sterile inflammation.

    PubMed

    Cai, Lin; Oyeniran, Clement; Biswas, Debolina D; Allegood, Jeremy; Milstien, Sheldon; Kordula, Tomasz; Maceyka, Michael; Spiegel, Sarah

    2016-08-01

    The bioactive sphingolipid metabolite, ceramide, regulates physiological processes important for inflammation and elevated levels of ceramide have been implicated in IL-1-mediated events. Although much has been learned about ceramide generation by activation of sphingomyelinases in response to IL-1, the contribution of the de novo pathway is not completely understood. Because yeast ORM1 and ORM2 proteins negatively regulate ceramide levels through inhibition of serine palmitoyltransferase, the first committed step in ceramide biosynthesis, we examined the functions of individual mammalian ORM orthologs, ORM (yeast)-like (ORMDL)1-3, in regulation of ceramide levels. In HepG2 liver cells, downregulation of ORMDL3 markedly increased the ceramide precursors, dihydrosphingosine and dihydroceramide, primarily from de novo biosynthesis based on [U-(13)C]palmitate incorporation into base-labeled and dual-labeled dihydroceramides, whereas downregulation of each isoform increased dihydroceramides [(13)C]labeled in only the amide-linked fatty acid. IL-1 and the IL-6 family cytokine, oncostatin M, increased dihydroceramide and ceramide levels in HepG2 cells and concomitantly decreased ORMDL proteins. Moreover, during irritant-induced sterile inflammation in mice leading to induction of the acute-phase response, which is dependent on IL-1, expression of ORMDL proteins in the liver was strongly downregulated and accompanied by increased ceramide levels in the liver and accumulation in the blood. Together, our results suggest that ORMDLs may be involved in regulation of ceramides during IL-1-mediated sterile inflammation. PMID:27313060

  4. Site specific incorporation of keto amino acids into proteins

    DOEpatents

    Schultz, Peter G.; Wang, Lei

    2009-04-28

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  5. Site specific incorporation of keto amino acids into proteins

    DOEpatents

    Schultz, Peter G.; Wang, Lei

    2008-10-07

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  6. Site specific incorporation of keto amino acids into proteins

    SciTech Connect

    Schultz, Peter G.; Wang, Lei

    2012-02-14

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  7. Site specific incorporation of keto amino acids into proteins

    SciTech Connect

    Schultz, Peter G.; Wang, Lei

    2011-12-06

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  8. Site specific incorporation of keto amino acids into proteins

    SciTech Connect

    Schultz, Peter G.; Wang, Lei

    2011-03-22

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  9. Effects of antinutritional factors on protein digestibility and amino acid availability in foods.

    PubMed

    Gilani, G Sarwar; Cockell, Kevin A; Sepehr, Estatira

    2005-01-01

    Digestibility of protein in traditional diets from developing countries such as India, Guatemala, and Brazil is considerably lower compared to that of protein in typical North American diets (54-78 versus 88-94%). The presence of less digestible protein fractions, high levels of insoluble fiber, and high concentrations of antinutritional factors in the diets of developing countries, which are based on less refined cereals and grain legumes as major sources of protein, are responsible for poor digestibility of protein. The effects of the presence of some of the important antinutritional factors on protein and amino digestibilities of food and feed products are reviewed in this chapter. Food and feed products may contain a number of antinutritional factors that may adversely affect protein digestibility and amino acid availability. Antinutritional factors may occur naturally, such as glucosinolates in mustard and rapeseed protein products, trypsin inhibitors and hemagglutinins in legumes, tannins in legumes and cereals, phytates in cereals and oilseeds, and gossypol in cottonseed protein products. Antinutritional factors may also be formed during heat/alkaline processing of protein products, yielding Maillard compounds, oxidized forms of sulfur amino acids, D-amino acids, and lysinoalanine (LAL, an unnatural amino acid derivative). The presence of high levels of dietary trypsin inhibitors from soybeans, kidney beans, or other grain legumes can cause substantial reductions in protein and amino acid digestibilities (up to 50%) in rats and pigs. Similarly, the presence of high levels of tannins in cereals, such as sorghum, and grain legumes, such as fababean (Vicia faba L.), can result in significantly reduced protein and amino acid digestibilities (up to 23%) in rats, poultry, and pigs. Studies involving phytase supplementation of production rations for swine or poultry have provided indirect evidence that normally encountered levels of phytates in cereals and legumes

  10. Does high serum uric acid level cause aspirin resistance?

    PubMed

    Yildiz, Bekir S; Ozkan, Emel; Esin, Fatma; Alihanoglu, Yusuf I; Ozkan, Hayrettin; Bilgin, Murat; Kilic, Ismail D; Ergin, Ahmet; Kaftan, Havane A; Evrengul, Harun

    2016-06-01

    In patients with coronary artery disease (CAD), though aspirin inhibits platelet activation and reduces atherothrombotic complications, it does not always sufficiently inhibit platelet function, thereby causing a clinical situation known as aspirin resistance. As hyperuricemia activates platelet turnover, aspirin resistance may be specifically induced by increased serum uric acid (SUA) levels. In this study, we thus investigated the association between SUA level and aspirin resistance in patients with CAD. We analyzed 245 consecutive patients with stable angina pectoris (SAP) who in coronary angiography showed more than 50% occlusion in a major coronary artery. According to aspirin resistance, two groups were formed: the aspirin resistance group (Group 1) and the aspirin-sensitive group (Group 2). Compared with those of Group 2, patients with aspirin resistance exhibited significantly higher white blood cell counts, neutrophil counts, neutrophil-to-lymphocyte ratios, SUA levels, high-sensitivity C-reactive protein levels, and fasting blood glucose levels. After multivariate analysis, a high level of SUA emerged as an independent predictor of aspirin resistance. The receiver-operating characteristic analysis provided a cutoff value of 6.45 mg/dl for SUA to predict aspirin resistance with 79% sensitivity and 65% specificity. Hyperuricemia may cause aspirin resistance in patients with CAD and high SUA levels may indicate aspirin-resistant patients. Such levels should thus recommend avoiding heart attack and stroke by adjusting aspirin dosage. PMID:26656902

  11. Phytic acid reduction in soy protein improves zinc bioavailability

    SciTech Connect

    Zhou, J.R.; Wong, M.S.; Burns, R.A.; Erdman, J.W. Jr. Mead Johnson Research Center, Evansville, IN )

    1991-03-15

    The objective of this study was to confirm previous studies that have suggested that reduction of phytic acid in soy improved zinc bioavailability (BV). Two commercially-produced soybean isolates containing either a normal phytic acid level or a reduced level were formulated into diets so as to provide 6 or 9 ppm zinc. Control diets were egg white protein-based and contained 3, 6 or 9 ppm zinc from zinc carbonate. Weanling male rats were fed these diets for 21 days and food intake and weight gain monitored. Slope ratio analysis of total tibia zinc content compared to total zinc intake revealed that zinc BV from reduced phytic acid soy isolate-containing diets was indistinguishable from control egg white diets. In contrast, zinc BV from normal soy isolate diets was significantly reduced compared to reduced phytic acid and control diets. These results coupled with other results indicate that phytic acid is the inhibitory factor in soybean products that results in reduced zinc BV.

  12. Regulation of intestinal protein metabolism by amino acids.

    PubMed

    Bertrand, Julien; Goichon, Alexis; Déchelotte, Pierre; Coëffier, Moïse

    2013-09-01

    Gut homeostasis plays a major role in health and may be regulated by quantitative and qualitative food intake. In the intestinal mucosa, an intense renewal of proteins occurs, at approximately 50% per day in humans. In some pathophysiological conditions, protein turnover is altered and may contribute to intestinal or systemic diseases. Amino acids are key effectors of gut protein turnover, both as constituents of proteins and as regulatory molecules limiting intestinal injury and maintaining intestinal functions. Many studies have focused on two amino acids: glutamine, known as the preferential substrate of rapidly dividing cells, and arginine, another conditionally essential amino acid. The effects of glutamine and arginine on protein synthesis appear to be model and condition dependent, as are the involved signaling pathways. The regulation of gut protein degradation by amino acids has been minimally documented until now. This review will examine recent data, helping to better understand how amino acids regulate intestinal protein metabolism, and will explore perspectives for future studies.

  13. Level of urinary liver-type fatty acid-binding protein is associated with cardiac markers and electrocardiographic abnormalities in type-2 diabetes with chronic kidney disease stage G1 and G2.

    PubMed

    Maeda, Yoshiteru; Suzuki, Atsushi; Ishii, Junnichi; Sekiguchi-Ueda, Sahoko; Shibata, Megumi; Yoshino, Yasumasa; Asano, Shogo; Hayakawa, Nobuki; Nakamura, Kazuhiro; Akiyama, Yasukazu; Kitagawa, Fumihiko; Sakuishi, Toshiaki; Fujita, Takashi; Hashimoto, Shuji; Ozaki, Yukio; Itoh, Mitsuyasu

    2015-05-01

    Urinary liver-type fatty acid-binding protein (L-FABP) reflects the degree of stress in proximal tubules of the kidney. We examined the level of L-FABP in type-2 diabetes mellitus (T2DM) patients with chronic kidney disease (CKD) stage G1 and G2, and its relationship with cardiac markers and electrocardiographic (ECG) abnormalities. T2DM patients whose estimated glomerular filtration rate (eGFR) was ≥60 mL/min/1.73 m(2) were recruited [n = 276 (165 males), mean age 64 years]. The median level of urinary L-FABP was 6.6 μg/gCr. Urinary L-FABP showed significant correlation with urinary albumin-to-creatinine ratio (ACR) (r = 0.51, p < 0.0001). Median (25th-75th percentile) eGFR was 82 (72-95) mL/min/1.73 m2. We divided patients into four subgroups (group 1, L-FABP ≤8.4 μg/gCr and ACR ≤30 mg/gCr; group 2, L-FABP ≤8.4 μg/gCr and ACR >30 mg/gCr; group 3, L-FABP >8.4 μg/gCr and ACR ≤30 mg/gCr; group 4, L-FABP >8.4 μg/gCr and ACR >30 mg/gCr). Compared with group 1, group 4 was significantly higher in systolic blood pressure, and eGFR using standardized serum cystatin C, high-sensitivity troponin T, and N-terminal pro-brain natriuretic peptide (NT-proBNP). Group 4 had significantly higher level of NT-proBNP than group 3. Groups 2, 3 and 4 showed more ECG abnormalities than group 1. These findings suggest that simultaneous measurement of urinary L-FABP and ACR should be useful to assess cardiovascular damage reflecting on the elevation of cardiac markers and ECG abnormalities in T2DM with CKD G1 and G2.

  14. Non-standard amino acid incorporation into proteins using Escherichia coli cell-free protein synthesis

    NASA Astrophysics Data System (ADS)

    Hong, Seok Hoon; Kwon, Yong-Chan; Jewett, Michael

    2014-06-01

    Incorporating non-standard amino acids (NSAAs) into proteins enables new chemical properties, new structures, and new functions. In recent years, improvements in cell-free protein synthesis (CFPS) systems have opened the way to accurate and efficient incorporation of NSAAs into proteins. The driving force behind this development has been three-fold. First, a technical renaissance has enabled high-yielding (>1 g/L) and long-lasting (>10 h in batch operation) CFPS in systems derived from Escherichia coli. Second, the efficiency of orthogonal translation systems has improved. Third, the open nature of the CFPS platform has brought about an unprecedented level of control and freedom of design. Here, we review recent developments in CFPS platforms designed to precisely incorporate NSAAs. In the coming years, we anticipate that CFPS systems will impact efforts to elucidate structure/function relationships of proteins and to make biomaterials and sequence-defined biopolymers for medical and industrial applications.

  15. Amino Acid Flux from Metabolic Network Benefits Protein Translation: the Role of Resource Availability

    PubMed Central

    Hu, Xiao-Pan; Yang, Yi; Ma, Bin-Guang

    2015-01-01

    Protein translation is a central step in gene expression and affected by many factors such as codon usage bias, mRNA folding energy and tRNA abundance. Despite intensive previous studies, how metabolic amino acid supply correlates with protein translation efficiency remains unknown. In this work, we estimated the amino acid flux from metabolic network for each protein in Escherichia coli and Saccharomyces cerevisiae by using Flux Balance Analysis. Integrated with the mRNA expression level, protein abundance and ribosome profiling data, we provided a detailed description of the role of amino acid supply in protein translation. Our results showed that amino acid supply positively correlates with translation efficiency and ribosome density. Moreover, with the rank-based regression model, we found that metabolic amino acid supply facilitates ribosome utilization. Based on the fact that the ribosome density change of well-amino-acid-supplied genes is smaller than poorly-amino-acid-supply genes under amino acid starvation, we reached the conclusion that amino acid supply may buffer ribosome density change against amino acid starvation and benefit maintaining a relatively stable translation environment. Our work provided new insights into the connection between metabolic amino acid supply and protein translation process by revealing a new regulation strategy that is dependent on resource availability. PMID:26056817

  16. Plasma Anti-Glial Fibrillary Acidic Protein Autoantibody Levels during the Acute and Chronic Phases of Traumatic Brain Injury: A Transforming Research and Clinical Knowledge in Traumatic Brain Injury Pilot Study.

    PubMed

    Wang, Kevin K W; Yang, Zhihui; Yue, John K; Zhang, Zhiqun; Winkler, Ethan A; Puccio, Ava M; Diaz-Arrastia, Ramon; Lingsma, Hester F; Yuh, Esther L; Mukherjee, Pratik; Valadka, Alex B; Gordon, Wayne A; Okonkwo, David O; Manley, Geoffrey T; Cooper, Shelly R; Dams-O'Connor, Kristen; Hricik, Allison J; Inoue, Tomoo; Maas, Andrew I R; Menon, David K; Schnyer, David M; Sinha, Tuhin K; Vassar, Mary J

    2016-07-01

    We described recently a subacute serum autoantibody response toward glial fibrillary acidic protein (GFAP) and its breakdown products 5-10 days after severe traumatic brain injury (TBI). Here, we expanded our anti-GFAP autoantibody (AutoAb[GFAP]) investigation to the multicenter observational study Transforming Research and Clinical Knowledge in TBI Pilot (TRACK-TBI Pilot) to cover the full spectrum of TBI (Glasgow Coma Scale 3-15) by using acute (<24 h) plasma samples from 196 patients with acute TBI admitted to three Level I trauma centers, and a second cohort of 21 participants with chronic TBI admitted to inpatient TBI rehabilitation. We find that acute patients self-reporting previous TBI with loss of consciousness (LOC) (n = 43) had higher day 1 AutoAb[GFAP] (mean ± standard error: 9.11 ± 1.42; n = 43) than healthy controls (2.90 ± 0.92; n = 16; p = 0.032) and acute patients reporting no previous TBI (2.97 ± 0.37; n = 106; p < 0.001), but not acute patients reporting previous TBI without LOC (8.01 ± 1.80; n = 47; p = 0.906). These data suggest that while exposure to TBI may trigger the AutoAb[GFAP] response, circulating antibodies are elevated specifically in acute TBI patients with a history of TBI. AutoAb[GFAP] levels for participants with chronic TBI (average post-TBI time 176 days or 6.21 months) were also significantly higher (15.08 ± 2.82; n = 21) than healthy controls (p < 0.001). These data suggest a persistent upregulation of the autoimmune response to specific brain antigen(s) in the subacute to chronic phase after TBI, as well as after repeated TBI insults. Hence, AutoAb[GFAP] may be a sensitive assay to study the dynamic interactions between post-injury brain and patient-specific autoimmune responses across acute and chronic settings after TBI.

  17. Atomic-level analysis of membrane-protein structure.

    PubMed

    Hendrickson, Wayne A

    2016-06-01

    Membrane proteins are substantially more challenging than natively soluble proteins as subjects for structural analysis. Thus, membrane proteins are greatly underrepresented in structural databases. Recently, focused consortium efforts and advances in methodology for protein production, crystallographic analysis and cryo-EM analysis have accelerated the pace of atomic-level structure determination of membrane proteins.

  18. High levels of vegetable oils in plant protein-rich diets fed to gilthead sea bream ( Sparus aurata L.): growth performance, muscle fatty acid profiles and histological alterations of target tissues.

    PubMed

    Benedito-Palos, Laura; Navarro, Juan C; Sitjà-Bobadilla, Ariadna; Bell, J Gordon; Kaushik, Sadasivam; Pérez-Sánchez, Jaume

    2008-11-01

    The feasibility of fish oil (FO) replacement by vegetable oils (VO) was investigated in gilthead sea bream (Sparus aurata L.) in a growth trial conducted for the duration of 8 months. Four isolipidic and isoproteic diets rich in plant proteins were supplemented with L-lysine (0.55 %) and soya lecithin (1 %). Added oil was either FO (control) or a blend of VO, replacing 33 % (33VO diet), 66 % (66VO diet) and 100 % (VO diet) of FO. No detrimental effects on growth performance were found with the partial FO replacement, but feed intake and growth rates were reduced by about 10 % in fish fed the VO diet. The replacement strategy did not damage the intestinal epithelium, and massive accumulation of lipid droplets was not found within enterocytes. All fish showed fatty livers, but signs of lipoid liver disease were only found in fish fed the VO diet. Muscle fatty acid profiles of total lipids reflected the diet composition with a selective incorporation of unsaturated fatty acids in polar lipids. The robustness of the phospholipid fatty acid profile when essential fatty acid requirements were theoretically covered by the diet was evidenced by multivariate principal components analysis in fish fed control, 33VO and 66VO diets.

  19. Expression of the whey acidic protein in transgenic pigs impairs mammary development.

    PubMed

    Shamay, A; Pursel, V G; Wilkinson, E; Wall, R J; Hennighausen, L

    1992-05-01

    The whey acidic protein has been found in milk of mice, rats, rabbits and camels, and its gene is expressed specifically in mammary tissue at late pregnancy and throughout lactation. A characteristic of whey acidic protein is the 'four-disulfide-core' signature which is also present in proteins involved in organ development. We have generated six lines of transgenic pigs which carry a mouse whey acidic protein transgene and express it at high levels in their mammary glands. Transgenic sows from three lines could not produce sufficient quantities of milk to support normal development of healthy offspring. This phenotype appears to be similar, if not identical, to the milchlos phenotype exhibited by mice expressing whey acidic protein transgenes. Mammary tissue from post-partum milchlos sows had an immature histological appearance, which was distinct from that observed during normal development or involution. Expression of the whey acidic protein transgene was found in mammary tissue from sexually immature pigs from milchlos lines, but not in sows from lines that appeared to lactate normally. We suggest that precocious synthesis of whey acidic protein impairs mammary development and function. Impaired mammary development due to inappropriate timing of whey acidic protein expression is consistent with the notion that proteins with the 'four-disulfide-core' signature participate in tissue formation. PMID:1284481

  20. Modelling and Characterization of Glial Fibrillary Acidic Protein

    PubMed Central

    Deka, Hemchandra; Sarmah, Rajeev; Sharma, Ankita; Biswas, Sagarika

    2015-01-01

    Glial Fibrillary Acidic Protein (GFAP) is an intermediate-filament (IF) protein that maintains the astrocytes of the Central Nervous System in Human. This is differentially expressed during serological studies in inflamed condition such as Rheumatoid Arthritis (RA). Therefore, it is of interest to glean molecular insight using a model of GFAP (49.88 kDa) due to its crystallographic nonavailability. The present study has been taken into consideration to construct computational protein model using Modeller 9.11. The structural relevance of the protein was verified using Gromacs 4.5 followed by validation through PROCHECK, Verify 3D, WHAT-IF, ERRAT and PROVE for reliability. The constructed three dimensional (3D) model of GFAP protein had been scrutinized to reveal the associated functions by identifying ligand binding sites and active sites. Molecular level interaction study revealed five possible surface cavities as active sites. The model finds application in further computational analysis towards drug discovery in order to minimize the effect of inflammation. PMID:26420920

  1. Retinoic acid binding protein in normal and neopolastic rat prostate.

    PubMed

    Gesell, M S; Brandes, M J; Arnold, E A; Isaacs, J T; Ueda, H; Millan, J C; Brandes, D

    1982-01-01

    Sucrose density gradient analysis of cytosol from normal and neoplastic rat prostatic tissues exhibited a peak of (3H) retinoic acid binding in the 2S region, corresponding to the cytoplasmic retinoic acid binding protein (cRABP). In the Fisher-Copenhagen F1 rat, cRABP was present in the lateral lobe, but could not be detected in the ventral nor in the dorsal prostatic lobes. Four sublines of the R-3327 rat prostatic tumor contained similar levels of this binding protein. The absence of cRABP in the normal tissue of origin of the R-3327 tumor, the rat dorsal prostate, and reappearance in the neoplastic tissues follows a pattern described in other human and animal tumors. The occurrence of cRABP in the well-differentiated as well as in the anaplastic R-3327 tumors in which markers which reflect a state of differentiation and hormonal regulation, such as androgen receptor, 5 alpha reductase, and secretory acid phosphatase are either markedly reduced or absent, points to cRABP as a marker of malignant transformation.

  2. Retinoic acid binding protein in normal and neopolastic rat prostate.

    PubMed

    Gesell, M S; Brandes, M J; Arnold, E A; Isaacs, J T; Ueda, H; Millan, J C; Brandes, D

    1982-01-01

    Sucrose density gradient analysis of cytosol from normal and neoplastic rat prostatic tissues exhibited a peak of (3H) retinoic acid binding in the 2S region, corresponding to the cytoplasmic retinoic acid binding protein (cRABP). In the Fisher-Copenhagen F1 rat, cRABP was present in the lateral lobe, but could not be detected in the ventral nor in the dorsal prostatic lobes. Four sublines of the R-3327 rat prostatic tumor contained similar levels of this binding protein. The absence of cRABP in the normal tissue of origin of the R-3327 tumor, the rat dorsal prostate, and reappearance in the neoplastic tissues follows a pattern described in other human and animal tumors. The occurrence of cRABP in the well-differentiated as well as in the anaplastic R-3327 tumors in which markers which reflect a state of differentiation and hormonal regulation, such as androgen receptor, 5 alpha reductase, and secretory acid phosphatase are either markedly reduced or absent, points to cRABP as a marker of malignant transformation. PMID:6283503

  3. Echinococcus granulosus fatty acid binding proteins subcellular localization.

    PubMed

    Alvite, Gabriela; Esteves, Adriana

    2016-05-01

    Two fatty acid binding proteins, EgFABP1 and EgFABP2, were isolated from the parasitic platyhelminth Echinococcus granulosus. These proteins bind fatty acids and have particular relevance in flatworms since de novo fatty acids synthesis is absent. Therefore platyhelminthes depend on the capture and intracellular distribution of host's lipids and fatty acid binding proteins could participate in lipid distribution. To elucidate EgFABP's roles, we investigated their intracellular distribution in the larval stage by a proteomic approach. Our results demonstrated the presence of EgFABP1 isoforms in cytosolic, nuclear, mitochondrial and microsomal fractions, suggesting that these molecules could be involved in several cellular processes.

  4. Physical activity as a determinant of fecal bile acid levels

    PubMed Central

    Wertheim, Betsy C.; Martínez, María Elena; Ashbeck, Erin L.; Roe, Denise J.; Jacobs, Elizabeth T.; Alberts, David S.; Thompson, Patricia A.

    2009-01-01

    Physical activity is protective against colon cancer, whereas colonic bile acid exposure is a suspected risk factor. While likely related, the association between physical activity and bile acid levels has not been well studied. Furthermore, the effect of triglycerides, which are known to modify bile acid levels, on this relationship has not been investigated. We conducted a cross-sectional analysis of baseline fecal bile acid levels for 735 colorectal adenoma formers obtained from participants in a phase III ursodeoxycholic acid chemoprevention trial. Compared to the lowest quartile of recreational physical activity duration, the highest quartile was associated with a 17% lower fecal bile acid concentration, adjusted for age, sex, dietary fiber intake, and body mass index (P = 0.042). Furthermore, consistent with a previously established relationship between serum triglyceride levels and bile acid metabolism, we stratified by triglyceride level and observed a 34% lower fecal bile acid concentration (highest versus lowest quartiles of physical activity) in individuals with low triglycerides (< 136 mg/dL; P = 0.002). In contrast, no association between physical activity and fecal bile acid concentration was observed for subjects with high triglycerides (≥ 136 mg/dL). Our results suggest that the biological mechanism responsible for the protective effect of physical activity on the incidence of colon cancer may be partially mediated by decreasing colonic bile acid exposure. However, this effect may be limited to individuals with lower triglyceride levels. PMID:19383885

  5. Amino acid sequences of proteins from Leptospira serovar pomona.

    PubMed

    Alves, S F; Lefebvre, R B; Probert, W

    2000-01-01

    This report describes a partial amino acid sequences from three putative outer envelope proteins from Leptospira serovar pomona. In order to obtain internal fragments for protein sequencing, enzymatic and chemical digestion was performed. The enzyme clostripain was used to digest the proteins 32 and 45 kDa. In situ digestion of 40 kDa molecular weight protein was accomplished using cyanogen bromide. The 32 kDa protein generated two fragments, one of 21 kDa and another of 10 kDa that yielded five residues. A fragment of 24 kDa that yielded nineteen residues of amino acids was obtained from 45 kDa protein. A fragment with a molecular weight of 20 kDa, yielding a twenty amino acids sequence from the 40 kDa protein.

  6. Protein quality of supplements and meal replacements. Amino acids and calculated indicators of protein quality.

    PubMed

    Marable, N L; Hinners, M L; Hardison, N W; Kehrberg, N L

    1980-09-01

    The amino acid composition of several types of dietary supplements and meal replacements was measured and compared with label values when available and to published values for egg. Calculated indicators of protein quality, such as chemical score, protein calorie:total calorie ratio, individual essential amino acid:total essential amino acid ratio, and total essential amino acid:total amino acid ratio were also compared for products, egg, and the estimated pattern of adult requirements. Predigested liquid protein products were notably lower in protein quality than other products. All non-predigested products compared favorably with egg in terms of protein quality, but were more expensive and had no advantages over regular meals in terms of protein quality as reducing aids or protein supplements.

  7. Protein and amino acid metabolism and requirements

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are the major structural and functional components of all cells of the body. Enzymes, membrane carriers, blood transport molecules, intracellular matrix, and even hair and fingernails are proteins, as are many hormones. Proteins also constitute a major portion of all membranes, and the cons...

  8. Phosphate acceptor amino acid residues in structural proteins of rhabdoviruses.

    PubMed

    Sokol, F; Tan, K B; McFalls, M L; Madore, P

    1974-07-01

    Partial acid hydrolysates of the [(32)P]phosphate- or [(3)H]serine-labeled proteins of purified vesicular stomatitis, rabies, Lagos bat, Mokola, or spring viremia of carp virions and of purified intracellular nucleocapsids of these viruses have been analyzed by paper electrophoresis for the presence of phosphorylated amino acids. Both phosphoserine and phosphothreonine, with the former predominant, were present in virion and nucleocapsid preparations that contained phosphoproteins. An exception was the fish rhabdovirus, which contained only phosphoserine. When vesicular stomatitis or rabies virus proteins were phosphorylated in a cell-free system by the virion-associated protein kinase and analyzed for the presence of phosphorylated amino acid residues, phosphoserine was again found to be more abundant than phosphothreonine. After in vitro protein phosphorylation, another phospho-compound, possibly a third phosphoamino acid, was detected in the partial acid hydrolysates of these viruses. PMID:4365328

  9. Interactive Effects of Indigestible Carbohydrates, Protein Type, and Protein Level on Biomarkers of Large Intestine Health in Rats.

    PubMed

    Taciak, Marcin; Barszcz, Marcin; Tuśnio, Anna; Pastuszewska, Barbara

    2015-01-01

    The effects of indigestible carbohydrates, protein type, and protein level on large intestine health were examined in rats. For 21 days, 12 groups of six 12-week-old male Wistar rats were fed diets with casein (CAS), or potato protein concentrate (PPC), providing 14% (lower protein level; LP), or 20% (higher protein level; HP) protein, and containing cellulose, resistant potato starch, or pectin. Fermentation end-products, pH, and β-glucuronidase levels in cecal digesta, and ammonia levels in colonic digesta were determined. Cecal digesta, tissue weights, cecal and colon morphology, and colonocyte DNA damage were also analyzed. Digesta pH was lower, whereas relative mass of cecal tissue and digesta were higher in rats fed pectin diets than in those fed cellulose. Cecal parameters were greater in rats fed PPC and HP diets than in those fed CAS and LP diets, respectively. Short-chain fatty acid (SCFA) concentrations were unaffected by protein or carbohydrate type. Total SCFA, acetic acid, and propionic acid concentrations were greater in rats fed LP diets than in those fed HP. Cecal pool of isobutyric and isovaleric acids was greater in rats fed PPC than in those fed CAS diets. PPC diets decreased phenol concentration and increased ammonia concentration in cecal and colonic digesta, respectively. Cecal crypt depth was greater in rats fed PPC and HP diets, and was unaffected by carbohydrates; whereas colonic crypt depth was greater in rats fed cellulose. Myenteron thickness in the cecum was unaffected by nutrition, but was greater in the colon of rats fed cellulose. Colonocyte DNA damage was greater in rats fed LP diets than in those fed HP diets, and was unaffected by carbohydrate or protein type. It was found that nutritional factors decreasing cecal digesta weight contribute to greater phenol production, increased DNA damage, and reduced ammonia concentration in the colon. PMID:26536028

  10. Interactive Effects of Indigestible Carbohydrates, Protein Type, and Protein Level on Biomarkers of Large Intestine Health in Rats

    PubMed Central

    Taciak, Marcin; Barszcz, Marcin; Tuśnio, Anna; Pastuszewska, Barbara

    2015-01-01

    The effects of indigestible carbohydrates, protein type, and protein level on large intestine health were examined in rats. For 21 days, 12 groups of six 12-week-old male Wistar rats were fed diets with casein (CAS), or potato protein concentrate (PPC), providing 14% (lower protein level; LP), or 20% (higher protein level; HP) protein, and containing cellulose, resistant potato starch, or pectin. Fermentation end-products, pH, and β-glucuronidase levels in cecal digesta, and ammonia levels in colonic digesta were determined. Cecal digesta, tissue weights, cecal and colon morphology, and colonocyte DNA damage were also analyzed. Digesta pH was lower, whereas relative mass of cecal tissue and digesta were higher in rats fed pectin diets than in those fed cellulose. Cecal parameters were greater in rats fed PPC and HP diets than in those fed CAS and LP diets, respectively. Short-chain fatty acid (SCFA) concentrations were unaffected by protein or carbohydrate type. Total SCFA, acetic acid, and propionic acid concentrations were greater in rats fed LP diets than in those fed HP. Cecal pool of isobutyric and isovaleric acids was greater in rats fed PPC than in those fed CAS diets. PPC diets decreased phenol concentration and increased ammonia concentration in cecal and colonic digesta, respectively. Cecal crypt depth was greater in rats fed PPC and HP diets, and was unaffected by carbohydrates; whereas colonic crypt depth was greater in rats fed cellulose. Myenteron thickness in the cecum was unaffected by nutrition, but was greater in the colon of rats fed cellulose. Colonocyte DNA damage was greater in rats fed LP diets than in those fed HP diets, and was unaffected by carbohydrate or protein type. It was found that nutritional factors decreasing cecal digesta weight contribute to greater phenol production, increased DNA damage, and reduced ammonia concentration in the colon. PMID:26536028

  11. Measuring protein-protein and protein-nucleic Acid interactions by biolayer interferometry.

    PubMed

    Sultana, Azmiri; Lee, Jeffrey E

    2015-01-01

    Biolayer interferometry (BLI) is a simple, optical dip-and-read system useful for measuring interactions between proteins, peptides, nucleic acids, small molecules, and/or lipids in real time. In BLI, a biomolecular bait is immobilized on a matrix at the tip of a fiber-optic sensor. The binding between the immobilized ligand and another molecule in an analyte solution produces a change in optical thickness at the tip and results in a wavelength shift proportional to binding. BLI provides direct binding affinities and rates of association and dissociation. This unit describes an efficient approach using streptavidin-based BLI to analyze DNA-protein and protein-protein interactions. A quantitative set of equilibrium binding affinities (K(d)) and rates of association and dissociation (k(a)/k(d)) can be measured in minutes using nanomole quantities of sample.

  12. Optimizing Scoring Function of Protein-Nucleic Acid Interactions with Both Affinity and Specificity

    PubMed Central

    Yan, Zhiqiang; Wang, Jin

    2013-01-01

    Protein-nucleic acid (protein-DNA and protein-RNA) recognition is fundamental to the regulation of gene expression. Determination of the structures of the protein-nucleic acid recognition and insight into their interactions at molecular level are vital to understanding the regulation function. Recently, quantitative computational approach has been becoming an alternative of experimental technique for predicting the structures and interactions of biomolecular recognition. However, the progress of protein-nucleic acid structure prediction, especially protein-RNA, is far behind that of the protein-ligand and protein-protein structure predictions due to the lack of reliable and accurate scoring function for quantifying the protein-nucleic acid interactions. In this work, we developed an accurate scoring function (named as SPA-PN, SPecificity and Affinity of the Protein-Nucleic acid interactions) for protein-nucleic acid interactions by incorporating both the specificity and affinity into the optimization strategy. Specificity and affinity are two requirements of highly efficient and specific biomolecular recognition. Previous quantitative descriptions of the biomolecular interactions considered the affinity, but often ignored the specificity owing to the challenge of specificity quantification. We applied our concept of intrinsic specificity to connect the conventional specificity, which circumvents the challenge of specificity quantification. In addition to the affinity optimization, we incorporated the quantified intrinsic specificity into the optimization strategy of SPA-PN. The testing results and comparisons with other scoring functions validated that SPA-PN performs well on both the prediction of binding affinity and identification of native conformation. In terms of its performance, SPA-PN can be widely used to predict the protein-nucleic acid structures and quantify their interactions. PMID:24098651

  13. Interaction of milk whey protein with common phenolic acids

    NASA Astrophysics Data System (ADS)

    Zhang, Hao; Yu, Dandan; Sun, Jing; Guo, Huiyuan; Ding, Qingbo; Liu, Ruihai; Ren, Fazheng

    2014-01-01

    Phenolics-rich foods such as fruit juices and coffee are often consumed with milk. In this study, the interactions of α-lactalbumin and β-lactoglobulin with the phenolic acids (chlorogenic acid, caffeic acid, ferulic acid, and coumalic acid) were examined. Fluorescence, CD, and FTIR spectroscopies were used to analyze the binding modes, binding constants, and the effects of complexation on the conformation of whey protein. The results showed that binding constants of each whey protein-phenolic acid interaction ranged from 4 × 105 to 7 × 106 M-n and the number of binding sites n ranged from 1.28 ± 0.13 to 1.54 ± 0.34. Because of these interactions, the conformation of whey protein was altered, with a significant reduction in the amount of α-helix and an increase in the amounts of β-sheet and turn structures.

  14. Comparison of Bile Acids and Acetaminophen Protein Adducts in Children and Adolescents with Acetaminophen Toxicity.

    PubMed

    James, Laura; Yan, Ke; Pence, Lisa; Simpson, Pippa; Bhattacharyya, Sudeepa; Gill, Pritmohinder; Letzig, Lynda; Kearns, Gregory; Beger, Richard

    2015-01-01

    Metabolomics approaches have enabled the study of new mechanisms of liver injury in experimental models of drug toxicity. Disruption of bile acid homeostasis is a known mechanism of drug induced liver injury. The relationship of individual bile acids to indicators of oxidative drug metabolism (acetaminophen protein adducts) and liver injury was examined in children with acetaminophen overdose, hospitalized children with low dose exposure to acetaminophen, and children with no recent exposure to acetaminophen. Nine bile acids were quantified through targeted metabolomic analysis in the serum samples of the three groups. Bile acids were compared to serum levels of acetaminophen protein adducts and alanine aminotransferase. Glycodeoxycholic acid, taurodeoxycholic acid, and glycochenodeoxycholic acid were significantly increased in children with acetaminophen overdose compared to healthy controls. Among patients with acetaminophen overdose, bile acids were higher in subjects with acetaminophen protein adduct values > 1.0 nmol/mL and modest correlations were noted for three bile acids and acetaminophen protein adducts as follows: taurodeoxycholic acid (R=0.604; p<0.001), glycodeoxycholic acid (R=0.581; p<0.001), and glycochenodeoxycholic acid (R=0.571; p<0.001). Variability in bile acids was greater among hospitalized children receiving low doses of acetaminophen than in healthy children with no recent acetaminophen exposure. Compared to bile acids, acetaminophen protein adducts more accurately discriminated among children with acetaminophen overdose, children with low dose exposure to acetaminophen, and healthy control subjects. In children with acetaminophen overdose, elevations of conjugated bile acids were associated with specific indicators of acetaminophen metabolism and non-specific indicators of liver injury. PMID:26208104

  15. Comparison of Bile Acids and Acetaminophen Protein Adducts in Children and Adolescents with Acetaminophen Toxicity.

    PubMed

    James, Laura; Yan, Ke; Pence, Lisa; Simpson, Pippa; Bhattacharyya, Sudeepa; Gill, Pritmohinder; Letzig, Lynda; Kearns, Gregory; Beger, Richard

    2015-01-01

    Metabolomics approaches have enabled the study of new mechanisms of liver injury in experimental models of drug toxicity. Disruption of bile acid homeostasis is a known mechanism of drug induced liver injury. The relationship of individual bile acids to indicators of oxidative drug metabolism (acetaminophen protein adducts) and liver injury was examined in children with acetaminophen overdose, hospitalized children with low dose exposure to acetaminophen, and children with no recent exposure to acetaminophen. Nine bile acids were quantified through targeted metabolomic analysis in the serum samples of the three groups. Bile acids were compared to serum levels of acetaminophen protein adducts and alanine aminotransferase. Glycodeoxycholic acid, taurodeoxycholic acid, and glycochenodeoxycholic acid were significantly increased in children with acetaminophen overdose compared to healthy controls. Among patients with acetaminophen overdose, bile acids were higher in subjects with acetaminophen protein adduct values > 1.0 nmol/mL and modest correlations were noted for three bile acids and acetaminophen protein adducts as follows: taurodeoxycholic acid (R=0.604; p<0.001), glycodeoxycholic acid (R=0.581; p<0.001), and glycochenodeoxycholic acid (R=0.571; p<0.001). Variability in bile acids was greater among hospitalized children receiving low doses of acetaminophen than in healthy children with no recent acetaminophen exposure. Compared to bile acids, acetaminophen protein adducts more accurately discriminated among children with acetaminophen overdose, children with low dose exposure to acetaminophen, and healthy control subjects. In children with acetaminophen overdose, elevations of conjugated bile acids were associated with specific indicators of acetaminophen metabolism and non-specific indicators of liver injury.

  16. Automated protein hydrolysis delivering sample to a solid acid catalyst for amino acid analysis.

    PubMed

    Masuda, Akiko; Dohmae, Naoshi

    2010-11-01

    In this study, we developed an automatic protein hydrolysis system using strong cation-exchange resins as solid acid catalysts. Examining several kinds of inorganic solid acids and cation-exchange resins, we found that a few cation-exchange resins worked as acid catalysts for protein hydrolysis when heated in the presence of water. The most efficient resin yielded amounts of amino acids that were over 70% of those recovered after conventional hydrolysis with hydrochloric acid and resulted in amino acid compositions matching the theoretical values. The solid-acid hydrolysis was automated by packing the resin into columns, combining the columns with a high-performance liquid chromatography system, and heating them. The amino acids that constitute a protein can thereby be determined, minimizing contamination from the environment.

  17. Yeast prions: Paramutation at the protein level?

    PubMed

    Tuite, Mick F

    2015-08-01

    Prions are proteins that have the potential to refold into a novel conformation that templates the conversion of like molecules to the altered infectious form. In the yeast Saccharomyces cerevisiae, trans-generational epigenetic inheritance can be mediated by a number of structurally and functionally diverse prions. Prionogenesis can confer both loss-of-function and gain-of-function properties to the prion protein and this in turn can have a major impact on host phenotype, short-term adaptation and evolution of new traits. Prionogenesis shares a number of properties in common with paramutation and can be considered as a mitotically and meiotically heritable change in protein conformation induced by trans-interactions between homologous proteins. PMID:26386407

  18. Butachlor impact on protein, free amino acid and glutamine contents, and on activity levels of aminotransferases, glutamate dehydrogenase and glutamine synthetase in the fresh water snail, Pila globosa (Swainson).

    PubMed

    Rajyalakshmi, T; Srinivas, T; Swamy, K V; Mohan, P M

    1996-08-01

    Biochemical changes followed in the freshwater snail Pila globosa (Swainson) during exposure to sublethal concentrations of the herbicide butachlor (26.6 ppm) in the ambient medium, at 3,6,12,24 and 48 h intervals, were marked by a significant decrease in total and soluble proteins, and an increase in free amino acids in foot and hepatopancreas up to 12 h before gradually recovering. Aminotransferase activities and glutamine content decreased during the early periods of exposure, while glutamate dehydrogenase activity increased. After an initial elevation, glutamate synthetase activity decreased at later intervals. Maximum effect of butachlor on the enzymes was seen after 12 h exposure. The extent of increase or decrease in different parameters examined varied between the two tissues studied. These changes are discussed in relation to the toxic stress of butachlor.

  19. Acid-degradable polyurethane particles for protein-based vaccines

    PubMed Central

    Bachelder, Eric M.; Beaudette, Tristan T.; Broaders, Kyle E.; Paramonov, Sergey E.; Dashe, Jesse; Fréchet, Jean M. J.

    2009-01-01

    Acid-degradable particles containing a model protein antigen, ovalbumin, were prepared from a polyurethane with acetal moieties embedded throughout the polymer, and characterized by dynamic light scattering and transmission electron microscopy. The small molecule degradation by-product of the particles was synthesized and tested in vitro for toxicity indicating an LC50 of 12,500 μg/ml. A new liquid chromatography-mass spectrometry technique was developed to monitor the in vitro degradation of these particles. The degradation by-product inside RAW macrophages was at its highest level after 24 hours of culture and was efficiently exocytosed until it was no longer detectable after four days. When tested in vitro, these particles induced a substantial increase in the presentation of the immunodominant ovalbumin-derived peptide SIINFEKL in both macrophages and dendritic cells. In addition, vaccination with these particles generated a cytotoxic T-lymphocyte response that was superior to both free ovalbumin and particles made from an analogous but slower-degrading acid-labile polyurethane polymer. Overall, we present a fully degradable polymer system with non-toxic by-products, which may find use in various biomedical applications including protein-based vaccines. PMID:18710254

  20. Analysis of proteins responsive to acetic acid in Acetobacter: molecular mechanisms conferring acetic acid resistance in acetic acid bacteria.

    PubMed

    Nakano, Shigeru; Fukaya, Masahiro

    2008-06-30

    Acetic acid bacteria are used for industrial vinegar production because of their remarkable ability to oxidize ethanol and high resistance to acetic acid. Although several molecular machineries responsible for acetic acid resistance in acetic acid bacteria have been reported, the entire mechanism that confers acetic acid resistance has not been completely understood. One of the promising methods to elucidate the entire mechanism is global analysis of proteins responsive to acetic acid by two-dimensional gel electrophoresis. Recently, two proteins whose production was greatly enhanced by acetic acid in Acetobacter aceti were identified to be aconitase and a putative ABC-transporter, respectively; furthermore, overexpression or disruption of the genes encoding these proteins affected acetic acid resistance in A. aceti, indicating that these proteins are involved in acetic acid resistance. Overexpression of each gene increased acetic acid resistance in Acetobacter, which resulted in an improvement in the productivity of acetic acid fermentation. Taken together, the results of the proteomic analysis and those of previous studies indicate that acetic acid resistance in acetic acid bacteria is conferred by several mechanisms. These findings also provide a clue to breed a strain having high resistance to acetic acid for vinegar fermentation.

  1. Requirement for the heart-type fatty acid binding protein in cardiac fatty acid utilization.

    PubMed

    Binas, B; Danneberg, H; McWhir, J; Mullins, L; Clark, A J

    1999-05-01

    Nonenzymatic cytosolic fatty acid binding proteins (FABPs) are abundantly expressed in many animal tissues with high rates of fatty acid metabolism. No physiological role has been demonstrated for any FABP, although these proteins have been implicated in transport of free long-chain fatty acids (LCFAs) and protection against LCFA toxicity. We report here that mice lacking heart-type FABP (H-FABP) exhibit a severe defect of peripheral (nonhepatic, non-fat) LCFA utilization. In these mice, the heart is unable to efficiently take up plasma LCFAs, which are normally its main fuel, and switches to glucose usage. Altered plasma levels of LCFAs, glucose, lactate and beta-hydroxybutyrate are consistent with depressed peripheral LCFA utilization, intensified carbohydrate usage, and increased hepatic LCFA oxidation; these changes are most pronounced under conditions favoring LCFA oxidation. H-FABP deficiency is only incompletely compensated, however, causing acute exercise intolerance and, at old age, a localized cardiac hypertrophy. These data establish a requirement for H-FABP in cardiac intracellular lipid transport and fuel selection and a major role in metabolic homeostasis. This new animal model should be particularly useful for investigating the significance of peripheral LCFA utilization for heart function, insulin sensitivity, and blood pressure.

  2. Refsum disease diagnostic marker phytanic acid alters the physical state of membrane proteins of liver mitochondria.

    PubMed

    Schönfeld, P; Struy, H

    1999-08-27

    Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid), a branched chain fatty acid accumulating in Refsum disease to high levels throughout the body, induces uncoupling of rat liver mitochondria similar to non-branched fatty acids (e.g. palmitic acid), but the contribution of the ADP/ATP carrier or the aspartate/glutamate carrier in phytanic acid-induced uncoupling is of minor importance. Possible deleterious effects of phytanic acid on membrane-linked energy coupling processes were studied by ESR spectroscopy using rat liver mitochondria and a membrane preparation labeled with the lipid-specific spin probe 5-doxylstearic acid (5-DSA) or the protein-specific spin probe MAL-TEMPO (4-maleimido-2,2,6, 6-tetramethyl-piperidine-1-oxyl). The effects of phytanic acid on phospholipid molecular dynamics and on the physical state of membrane proteins were quantified by estimation of the order parameter or the ratio of the amplitudes of the weakly to strongly immobilized MAL-TEMPO binding sites (W/S ratio), respectively. It was found, that phytanic acid (1) increased the mobility of phospholipid molecules (indicated by a decrease in the order parameter) and (2) altered the conformational state and/or the segmental mobility of membrane proteins (indicated by a drastic decrease in the W/S ratio). Unsaturated fatty acids with multiple cis-double bonds (e.g. linolenic or arachidonic acid), but not non-branched FFA (ranging from chain length C10:0 to C18:0), also decrease the W/S ratio. It is hypothesized that the interaction of phytanic acid with transmembrane proteins might stimulate the proton permeability through the mitochondrial inner membrane according to a mechanism, different to a protein-supported fatty acid cycling.

  3. Serum Uric Acid Level among Acute Stroke Patients.

    PubMed

    Mohsin, M; Das, S N; Haque, M F; Shikha, S S; Bhuiyan, A S; Saha, P L; Das, B R; Chowdhury, M; Jahan, M K; Rahman, M M

    2016-04-01

    Stroke is one of the leading causes of death and disability in developed as well as developing countries like Bangladesh. Elevated serum uric acid levels may predict an increased risk for cerebro-vascular (CV) events including stroke. Aim of the study was to measure the serum uric acid level among stroke patients and determine the relationship between serum uric acid level and stroke. This descriptive, cross-sectional study was carried out in Department of Medicine, Mymensingh Medical College Hospital, Mymensingh, Bangladesh to measure serum uric acid level among 102 stroke patients in a period of one year by using non-probability sampling procedure. Finally, collected data were analyzed using SPSS software Version 17.0. It was observed that the mean age of patients was 60.87±8.05 years, of them 80(78.43%) patients were male and the rest 22(21.57%) were female. About 66(64.70%) of respondents were in age group 60 years and above, while 36(35.30%) were in age group 59 years and below. At least 23(22.55%) of stroke patients had elevated serum uric acid with a mean serum uric acid level of 5.18mg/dl and standard deviation 1.26mg/dl. About 23(27.38%) patients in ischemic stroke had elevated serum uric acid whereas 18(100%) patients in hemorrhagic stroke had normal uric acid level. Uric acid level was elevated in ischemic stroke than haemorrhagic stroke patients (p<0.001). High uric acid level may be considered as a risk factor in patients with acute ischemic stroke. PMID:27277350

  4. Protein Consumption and the Elderly: What Is the Optimal Level of Intake?

    PubMed Central

    Baum, Jamie I.; Kim, Il-Young; Wolfe, Robert R.

    2016-01-01

    Maintaining independence, quality of life, and health is crucial for elderly adults. One of the major threats to living independently is the loss of muscle mass, strength, and function that progressively occurs with aging, known as sarcopenia. Several studies have identified protein (especially the essential amino acids) as a key nutrient for muscle health in elderly adults. Elderly adults are less responsive to the anabolic stimulus of low doses of amino acid intake compared to younger individuals. However, this lack of responsiveness in elderly adults can be overcome with higher levels of protein (or essential amino acid) consumption. The requirement for a larger dose of protein to generate responses in elderly adults similar to the responses in younger adults provides the support for a beneficial effect of increased protein in older populations. The purpose of this review is to present the current evidence related to dietary protein intake and muscle health in elderly adults. PMID:27338461

  5. Advantages and limitations of the protein digestibility-corrected amino acid score (PDCAAS) as a method for evaluating protein quality in human diets.

    PubMed

    Schaafsma, Gertjan

    2012-08-01

    PDCAAS is a widely used assay for evaluating protein quality. It is a chemical score, which is derived from the ratio between the first limiting amino acid in a test protein and the corresponding amino acid in a reference amino acid pattern and corrected for true faecal N digestibility. Chemical scores exceeding 100 % are truncated to 100 %. The advantages of the PDCAAS are its simplicity and direct relationship to human protein requirements. The limitations are as follows: the reference pattern is based on the minimum amino acid requirements for tissue growth and maintenance and does not necessarily reflect the optimum intake. Truncated PDCAAS of high-quality proteins do not give any information about the power of these proteins to compensate, as a supplement, for low levels of dietary essential amino acids in low-quality proteins. It is likely that faecal N digestibility does not take into account the loss from the colon of indispensable amino acids that were not absorbed in the ileum. Anti-nutritional factors, such as lectins and trypsin inhibitors, in several plant protein sources can cause heightened endogenous losses of amino acids, an issue which is particularly relevant in animal feedstuffs. The assumption that amino acid supplementation can completely restore biological efficiency of the protein source is incorrect since the kinetics of digestion and absorption between supplemented free amino acids and amino acids present in dietary proteins, are different. PMID:23107546

  6. Ethanol Effects On Physiological Retinoic Acid Levels

    PubMed Central

    Napoli, Joseph L.

    2011-01-01

    Summary All-trans-retinoic acid (atRA) serves essential functions during embryogenesis and throughout post-natal vertebrate life. Insufficient or excess atRA causes teratogenic and/or toxic effects in the developing embryo: interference with atRA biosynthesis or signaling likely underlies some forms of cancer. Many symptoms of vitamin A (atRA precursor) deficiency and/or toxicity overlap with those of another pleiotropic agent—ethanol. These overlapping symptoms have prompted research to understand whether interference with atRA biosynthesis and/or action may explain (in part) pathology associated with excess ethanol consumption. Ethanol affects many aspects of retinoid metabolism and mechanisms of action site-specifically, but no robust data support inhibition of vitamin A metabolism, resulting in decreased atRA in vivo during normal vitamin A nutriture. Actually, ethanol either has no effect on or increases atRA at select sites. Despite this realization, insight into whether interactions between ethanol and retinoids represent cause vs. effect requires additional research. PMID:21766417

  7. Differential Occurrence of Interactions and Interaction Domains in Proteins Containing Homopolymeric Amino Acid Repeats.

    PubMed

    Pelassa, Ilaria; Fiumara, Ferdinando

    2015-01-01

    Homopolymeric amino acids repeats (AARs), which are widespread in proteomes, have often been viewed simply as spacers between protein domains, or even as "junk" sequences with no obvious function but with a potential to cause harm upon expansion as in genetic diseases associated with polyglutamine or polyalanine expansions, including Huntington disease and cleidocranial dysplasia. A growing body of evidence indicates however that at least some AARs can form organized, functional protein structures, and can regulate protein function. In particular, certain AARs can mediate protein-protein interactions, either through homotypic AAR-AAR contacts or through heterotypic contacts with other protein domains. It is still unclear however, whether AARs may have a generalized, proteome-wide role in shaping protein-protein interaction networks. Therefore, we have undertaken here a bioinformatics screening of the human proteome and interactome in search of quantitative evidence of such a role. We first identified the sets of proteins that contain repeats of any one of the 20 amino acids, as well as control sets of proteins chosen at random in the proteome. We then analyzed the connectivity between the proteins of the AAR-containing protein sets and we compared it with that observed in the corresponding control networks. We find evidence for different degrees of connectivity in the different AAR-containing protein networks. Indeed, networks of proteins containing polyglutamine, polyglutamate, polyproline, and other AARs show significantly increased levels of connectivity, whereas networks containing polyleucine and other hydrophobic repeats show lower degrees of connectivity. Furthermore, we observed that numerous protein-protein, -nucleic acid, and -lipid interaction domains are significantly enriched in specific AAR protein groups. These findings support the notion of a generalized, combinatorial role of AARs, together with conventional protein interaction domains, in shaping

  8. [Serum uric acid levels and risk of developing preeclampsia].

    PubMed

    Corominas, Ana I; Balconi, Silvia M; Palermo, Mario; Maskin, Bernardo; Damiano, Alicia E

    2014-01-01

    It is well known that preeclampsia is associated to high uric acid levels, but the clinical assessment of this relationship is still under consideration. Our research was to evaluate if periodic doses of uric acid during pregnancy might help to identify a high risk group prior to the onset of preeclampsia. We conducted a retrospective investigation in 79 primary gestates with normal blood pressure and 79 women with preeclampsia who were assisted at Hospital Nacional Posadas during 2010. Serum uric acid levels, creatininemia, uremia, and proteinuria data from the clinical records of the pregnant women were considered. Uric acid levels were similar in both groups during the first half of gestation. However, as of the 20th week, uric acid increased 1.5-times in preeclamptic women with no changes in creatinine and urea, confirming that these patients had no renal complications. Furthermore, we noted that higher levels of uric acid correlated with low birth weight. We also observed that pregnant women with a family history of hypertension were more likely to develop this condition. Moreover, we did not find a direct relationship with the fetal sex or the appearance of clinical symptoms. The analytical evidence suggests that changes in uric acid concentrations may be due to metabolic alterations at the initial stages of preeclampsia. Therefore, we propose that monitoring levels of uric acid during pregnancy might contribute to the early control of this condition.

  9. Salivary Sialic Acid Levels in Smokeless Tobacco Users

    PubMed Central

    Farhad Mollashahi, Leila; Honarmand, Marieh; Nakhaee, Alireza; Mollashahi, Ghasem

    2016-01-01

    Background Smokeless tobacco chewing is one of the known risk factors for oral cancer. It is consumed widely by residents of southeastern Iran. Objectives In this study, salivary free and total sialic acid, and total protein were compared in paan consumers and non-consumers. Patients and Methods In this cross-sectional study, unstimulated saliva of 94 subjects (44 paan consumers and 50 non-consumers) who were referred to the oral medicine department of the dentistry school of Zahedan were collected. Salivary free and total sialic acid, and total protein concentration were measured by standard biochemical methods, and the obtained data were analyzed using SPSS 20 through the non-parametric Mann-Whitney test. Results The concentration of salivary free sialic acid (23.21 ± 18.98 mg/L) was significantly increased in paan consumers. The concentration of salivary Total sialic acid (TSA) (39.57 ± 26.58 mg/L) and total protein (0.77 ± 0.81 mg/mL) showed increases in paan consumers, however, the results were not statistically significant. Conclusions Salivary free and total sialic acid, and total protein were higher in the paan consumers compared to non-consumers. Due to the carcinogenic effect of smokeless tobacco, measurement of these parameters in saliva may be useful in early detection of oral cancer. PMID:27622172

  10. Salivary Sialic Acid Levels in Smokeless Tobacco Users

    PubMed Central

    Farhad Mollashahi, Leila; Honarmand, Marieh; Nakhaee, Alireza; Mollashahi, Ghasem

    2016-01-01

    Background Smokeless tobacco chewing is one of the known risk factors for oral cancer. It is consumed widely by residents of southeastern Iran. Objectives In this study, salivary free and total sialic acid, and total protein were compared in paan consumers and non-consumers. Patients and Methods In this cross-sectional study, unstimulated saliva of 94 subjects (44 paan consumers and 50 non-consumers) who were referred to the oral medicine department of the dentistry school of Zahedan were collected. Salivary free and total sialic acid, and total protein concentration were measured by standard biochemical methods, and the obtained data were analyzed using SPSS 20 through the non-parametric Mann-Whitney test. Results The concentration of salivary free sialic acid (23.21 ± 18.98 mg/L) was significantly increased in paan consumers. The concentration of salivary Total sialic acid (TSA) (39.57 ± 26.58 mg/L) and total protein (0.77 ± 0.81 mg/mL) showed increases in paan consumers, however, the results were not statistically significant. Conclusions Salivary free and total sialic acid, and total protein were higher in the paan consumers compared to non-consumers. Due to the carcinogenic effect of smokeless tobacco, measurement of these parameters in saliva may be useful in early detection of oral cancer.

  11. Bile salt recognition by human liver fatty acid binding protein.

    PubMed

    Favretto, Filippo; Santambrogio, Carlo; D'Onofrio, Mariapina; Molinari, Henriette; Grandori, Rita; Assfalg, Michael

    2015-04-01

    Fatty acid binding proteins (FABPs) act as intracellular carriers of lipid molecules, and play a role in global metabolism regulation. Liver FABP (L-FABP) is prominent among FABPs for its wide ligand repertoire, which includes long-chain fatty acids as well as bile acids (BAs). In this work, we performed a detailed molecular- and atomic-level analysis of the interactions established by human L-FABP with nine BAs to understand the binding specificity for this important class of cholesterol-derived metabolites. Protein-ligand complex formation was monitored using heteronuclear NMR, steady-state fluorescence spectroscopy, and mass spectrometry. BAs were found to interact with L-FABP with dissociation constants in the narrow range of 0.6-7 μm; however, the diverse substitution patterns of the sterol nucleus and the presence of side-chain conjugation resulted in complexes endowed with various degrees of conformational heterogeneity. Trihydroxylated BAs formed monomeric complexes in which single ligand molecules occupied similar internal binding sites, based on chemical-shift perturbation data. Analysis of NMR line shapes upon progressive addition of taurocholate indicated that the binding mechanism departed from a simple binary association equilibrium, and instead involved intermediates along the binding path. The co-linear chemical shift behavior observed for L-FABP complexes with cholate derivatives added insight into conformational dynamics in the presence of ligands. The observed spectroscopic features of L-FABP/BA complexes, discussed in relation to ligand chemistry, suggest possible molecular determinants of recognition, with implications regarding intracellular BA transport. Our findings suggest that human L-FABP is a poorly selective, universal BA binder. PMID:25639618

  12. Bile salt recognition by human liver fatty acid binding protein.

    PubMed

    Favretto, Filippo; Santambrogio, Carlo; D'Onofrio, Mariapina; Molinari, Henriette; Grandori, Rita; Assfalg, Michael

    2015-04-01

    Fatty acid binding proteins (FABPs) act as intracellular carriers of lipid molecules, and play a role in global metabolism regulation. Liver FABP (L-FABP) is prominent among FABPs for its wide ligand repertoire, which includes long-chain fatty acids as well as bile acids (BAs). In this work, we performed a detailed molecular- and atomic-level analysis of the interactions established by human L-FABP with nine BAs to understand the binding specificity for this important class of cholesterol-derived metabolites. Protein-ligand complex formation was monitored using heteronuclear NMR, steady-state fluorescence spectroscopy, and mass spectrometry. BAs were found to interact with L-FABP with dissociation constants in the narrow range of 0.6-7 μm; however, the diverse substitution patterns of the sterol nucleus and the presence of side-chain conjugation resulted in complexes endowed with various degrees of conformational heterogeneity. Trihydroxylated BAs formed monomeric complexes in which single ligand molecules occupied similar internal binding sites, based on chemical-shift perturbation data. Analysis of NMR line shapes upon progressive addition of taurocholate indicated that the binding mechanism departed from a simple binary association equilibrium, and instead involved intermediates along the binding path. The co-linear chemical shift behavior observed for L-FABP complexes with cholate derivatives added insight into conformational dynamics in the presence of ligands. The observed spectroscopic features of L-FABP/BA complexes, discussed in relation to ligand chemistry, suggest possible molecular determinants of recognition, with implications regarding intracellular BA transport. Our findings suggest that human L-FABP is a poorly selective, universal BA binder.

  13. Protein and amino acid metabolism in the human newborn.

    PubMed

    Kalhan, Satish C; Bier, Dennis M

    2008-01-01

    Birth and adaptation to extrauterine life involve major shifts in the protein and energy metabolism of the human newborn. These include a shift from a state of continuous supply of nutrients including amino acids from the mother to cyclic periodic oral intake, a change in the redox state of organs, thermogenesis, and a significant change in the mobilization and use of oxidative substrates. The development of safe, stable isotopic tracer methods has allowed the study of protein and amino acid metabolism not only in the healthy newborn but also in those born prematurely and of low birth weight. These studies have identified the unique and quantitative aspects of amino acid/protein metabolism in the neonate, thus contributing to rational nutritional care of these babies. The present review summarizes the contemporary data on some of the significant developments in essential and dispensable amino acids and their relationship to overall protein metabolism. Specifically, the recent data of kinetics of leucine, phenylalanine, glutamine, sulfur amino acid, and threonine and their relation to whole-body protein turnover are presented. Finally, the physiological rationale and the impact of nutrient (amino acids) interventions on the dynamics of protein metabolism are discussed.

  14. Fatty acid transfer between multilamellar liposomes and fatty acid-binding proteins.

    PubMed

    Brecher, P; Saouaf, R; Sugarman, J M; Eisenberg, D; LaRosa, K

    1984-11-10

    A simple experimental system was developed for studying the movement of long-chain fatty acids between multilamellar liposomes and soluble proteins capable of binding fatty acids. Oleic acid was incorporated into multilamellar liposomes containing cholesterol and egg yolk lecithin and incubated with albumin or hepatic fatty acid-binding protein. It was found that the fatty acid transferred from the liposomes to either protein rapidly and selectively under conditions where phospholipid and cholesterol transfer did not occur. More than 50% of the fatty acid contained within liposomes could become protein bound, suggesting that the fatty acid moved readily between and across phospholipid bilayers. Transfer was reduced at low pH, and this reduction appeared to result from decreased dissociation of the protonated fatty acid from the bilayer. Liposomes made with dimyristoyl or dipalmitoyl lecithin and containing 1 mol per cent palmitic acid were used to show the effect of temperature on fatty acid transfer. Transfer to either protein did not occur at temperatures where the liposomes were in a gel state but occurred rapidly at temperatures at or above the transition temperatures of the phospholipid used. PMID:6490659

  15. Fatty Acid-Binding Protein 5 Facilitates the Blood-Brain Barrier Transport of Docosahexaenoic Acid.

    PubMed

    Pan, Yijun; Scanlon, Martin J; Owada, Yuji; Yamamoto, Yui; Porter, Christopher J H; Nicolazzo, Joseph A

    2015-12-01

    The brain has a limited ability to synthesize the essential polyunsaturated fatty acid (PUFA) docosahexaenoic acid (DHA) from its omega-3 fatty acid precursors. Therefore, to maintain brain concentrations of this PUFA at physiological levels, plasma-derived DHA must be transported across the blood-brain barrier (BBB). While DHA is able to partition into the luminal membrane of brain endothelial cells, its low aqueous solubility likely limits its cytosolic transfer to the abluminal membrane, necessitating the requirement of an intracellular carrier protein to facilitate trafficking of this PUFA across the BBB. As the intracellular carrier protein fatty acid-binding protein 5 (FABP5) is expressed at the human BBB, the current study assessed the putative role of FABP5 in the brain endothelial cell uptake and BBB transport of DHA in vitro and in vivo, respectively. hFAPB5 was recombinantly expressed and purified from Escherichia coli C41(DE3) cells and the binding affinity of DHA to hFABP5 assessed using isothermal titration calorimetry. The impact of FABP5 siRNA on uptake of (14)C-DHA into immortalized human brain microvascular endothelial (hCMEC/D3) cells was assessed. An in situ transcardiac perfusion method was optimized in C57BL/6 mice and subsequently used to compare the BBB influx rate (Kin) of (14)C-DHA between FABP5-deficient (FABP5(-/-)) and wild-type (FABP5(+/+)) C57BL/6 mice. DHA bound to hFABP5 with an equilibrium dissociation constant of 155 ± 8 nM (mean ± SEM). FABP5 siRNA transfection decreased hCMEC/D3 mRNA and protein expression of FABP5 by 53.2 ± 5.5% and 44.8 ± 13.7%, respectively, which was associated with a 14.1 ± 2.7% reduction in (14)C-DHA cellular uptake. By using optimized conditions for the in situ transcardiac perfusion (a 1 min preperfusion (10 mL/min) followed by perfusion of (14)C-DHA (1 min)), the Kin of (14)C-DHA was 0.04 ± 0.01 mL/g/s. Relative to FABP5(+/+) mice, the Kin of (14)C-DHA decreased 36.7 ± 12.4% in FABP5(-/-) mice

  16. Conformational Entropy of Intrinsically Disordered Proteins from Amino Acid Triads

    PubMed Central

    Baruah, Anupaul; Rani, Pooja; Biswas, Parbati

    2015-01-01

    This work quantitatively characterizes intrinsic disorder in proteins in terms of sequence composition and backbone conformational entropy. Analysis of the normalized relative composition of the amino acid triads highlights a distinct boundary between globular and disordered proteins. The conformational entropy is calculated from the dihedral angles of the middle amino acid in the amino acid triad for the conformational ensemble of the globular, partially and completely disordered proteins relative to the non-redundant database. Both Monte Carlo (MC) and Molecular Dynamics (MD) simulations are used to characterize the conformational ensemble of the representative proteins of each group. The results show that the globular proteins span approximately half of the allowed conformational states in the Ramachandran space, while the amino acid triads in disordered proteins sample the entire range of the allowed dihedral angle space following Flory’s isolated-pair hypothesis. Therefore, only the sequence information in terms of the relative amino acid triad composition may be sufficient to predict protein disorder and the backbone conformational entropy, even in the absence of well-defined structure. The predicted entropies are found to agree with those calculated using mutual information expansion and the histogram method. PMID:26138206

  17. Conformational Entropy of Intrinsically Disordered Proteins from Amino Acid Triads.

    PubMed

    Baruah, Anupaul; Rani, Pooja; Biswas, Parbati

    2015-07-03

    This work quantitatively characterizes intrinsic disorder in proteins in terms of sequence composition and backbone conformational entropy. Analysis of the normalized relative composition of the amino acid triads highlights a distinct boundary between globular and disordered proteins. The conformational entropy is calculated from the dihedral angles of the middle amino acid in the amino acid triad for the conformational ensemble of the globular, partially and completely disordered proteins relative to the non-redundant database. Both Monte Carlo (MC) and Molecular Dynamics (MD) simulations are used to characterize the conformational ensemble of the representative proteins of each group. The results show that the globular proteins span approximately half of the allowed conformational states in the Ramachandran space, while the amino acid triads in disordered proteins sample the entire range of the allowed dihedral angle space following Flory's isolated-pair hypothesis. Therefore, only the sequence information in terms of the relative amino acid triad composition may be sufficient to predict protein disorder and the backbone conformational entropy, even in the absence of well-defined structure. The predicted entropies are found to agree with those calculated using mutual information expansion and the histogram method.

  18. HALOACETIC ACIDS PERTURB PROTEIN PHOSPHORYLATION IN MOUSE EMBRYOS IN VITRO

    EPA Science Inventory

    HALOACETIC ACIDS PERTURB PROTEIN PHOSPHORYLATION IN MOUSE EMBRYOS IN VITRO. MR Blanton and ES Hunter. Reproductive Toxicology Division, NHEERL, ORD, US EPA, RTP, NC, USA.
    Sponsor: JM Rogers.
    Haloacetic Acids (HAAs) formed during the disinfection process are present in drin...

  19. Glial fibrillary acidic protein as a biomarker for brain injury in neonatal CHD.

    PubMed

    McKenney, Stephanie L; Mansouri, Fahad F; Everett, Allen D; Graham, Ernest M; Burd, Irina; Sekar, Priya

    2016-10-01

    Neonates with critical CHD have evidence, by imaging, of preoperative brain injury, although the timing is unknown. We used circulating postnatal serum glial fibrillary acidic protein as a measure of acute perinatal brain injury in neonates with CHD. Glial fibrillary acidic protein was measured on admission and daily for the first 4 days of life in case and control groups; we included two control groups in this study - non-brain-injured newborns and brain-injured newborns. Comparisons were performed using the Kruskal-Wallis test with Dunn's multiple comparisons, Student's t-test, and χ2 test of independence where appropriate. In aggregate, there were no significant differences in overall glial fibrillary acidic protein levels between CHD patients (n=56) and negative controls (n=23) at any time point. By day 4 of life, 7/56 (12.5%) CHD versus 0/23 (0%) normal controls had detectable glial fibrillary acidic protein levels. Although not statistically significant, the 5/10 (50%) left heart obstruction group versus 1/17 (6%) conoventricular, 0/13 (0%) right heart, and 1/6 (17%) septal defect patients trended towards elevated levels of glial fibrillary acidic protein at day 4 of life. Overall, glial fibrillary acidic protein reflected no evidence for significant peripartum brain injury in neonates with CHD, but there was a trend for elevation by postnatal day 4 in neonates with left heart obstruction. This pilot study suggests that methods such as monitoring glial fibrillary acidic protein levels may provide new tools to optimise preoperative care and neuroprotection in high-risk neonates with specific types of CHD.

  20. Amyloid protein precursor stimulates excitatory amino acid transport. Implications for roles in neuroprotection and pathogenesis.

    PubMed

    Masliah, E; Raber, J; Alford, M; Mallory, M; Mattson, M P; Yang, D; Wong, D; Mucke, L

    1998-05-15

    Excitatory neurotransmitters such as glutamate are required for the normal functioning of the central nervous system but can trigger excitotoxic neuronal injury if allowed to accumulate to abnormally high levels. Their extracellular levels are controlled primarily by transmitter uptake into astrocytes. Here, we demonstrate that the amyloid protein precursor may participate in the regulation of this important process. The amyloid protein precursor has been well conserved through evolution, and a number of studies indicate that it may function as an endogenous excitoprotectant. However, the mechanisms underlying this neuroprotective capacity remain largely unknown. At moderate levels of expression, human amyloid protein precursors increased glutamate/aspartate uptake in brains of transgenic mice, with the 751-amino acid isoform showing greater potency than the 695-amino acid isoform. Cerebral glutamate/aspartate transporter protein levels were higher in transgenic mice than in non-transgenic controls, whereas transporter mRNA levels were unchanged. Amyloid protein precursor-dependent stimulation of aspartate uptake by cultured primary astrocytes was associated with increases in protein kinase A and C activity and could be blocked by inhibitors of these kinases. The stimulation of astroglial excitatory amino acid transport by amyloid protein precursors could protect the brain against excitotoxicity and may play an important role in neurotransmission. PMID:9575214

  1. Effect of maternal micronutrients (folic acid, vitamin B12) and omega 3 fatty acids on liver fatty acid desaturases and transport proteins in Wistar rats.

    PubMed

    Wadhwani, Nisha S; Manglekar, Rupali R; Dangat, Kamini D; Kulkarni, Asmita V; Joshi, Sadhana R

    2012-01-01

    A disturbed fatty acid metabolism increases the risk of adult non-communicable diseases. This study examines the effect of maternal micronutrients on the fatty acid composition, desaturase activity, mRNA levels of fatty acid desaturases and transport proteins in the liver. Pregnant female rats were divided into 6 groups at 2 levels of folic acid both in the presence and absence of vitamin B(12). The vitamin B(12) deficient groups were supplemented with omega 3 fatty acid. An imbalance of maternal micronutrients reduces liver docosahexaenoic acid, increases Δ5 desaturase activity but decreases mRNA levels, decreases Δ6 desaturase activity but not mRNA levels as compared to control. mRNA level of Δ5 desaturase reverts back to the levels of the control group as a result of omega 3 fatty acid supplementation. Our data for the first time indicates that maternal micronutrients differentially alter the activity and expression of fatty acid desaturases in the liver.

  2. Quality of buffalo milk as affected by dietary protein level and flaxseed supplementation.

    PubMed

    Santillo, A; Caroprese, M; Marino, R; Sevi, A; Albenzio, M

    2016-10-01

    The aim of the present research was to evaluate the effects of protein level and flaxseed supplementation on the yield and quality of buffalo milk. In particular, the fatty acid profile of milk from buffalo cows subjected to different diets has been investigated. A 2×3 factorial design was tested with buffalo cows receiving 2 dietary crude protein (CP) and 3 flaxseed (FS) supplementation levels. Treatments were (1) low dietary CP level [12% of dry matter (DM)] and no flaxseed supplementation (LP); (2) low dietary CP level (12% of DM) and low flaxseed supplementation (500g/d) (LPFS500); (3) low dietary CP level (12% of DM) and moderate flaxseed supplementation (1,000g/d) (LPFS1000); (4) moderate dietary CP level (15% of DM) and no flaxseed supplementation (MP); (5) moderate dietary CP level (15% of DM) and low flaxseed supplementation (500g/d) (MPFS500); and (6) moderate dietary CP level (15% of DM) and moderate flaxseed supplementation (1,000g/d) (MPFS1000). Milk protein and casein were affected by flaxseed supplementation being higher in MP, intermediate in LP, and lower in flaxseed-supplemented diets. However, the results from the present study highlighted that low protein diets sustained milk yield, protein, and casein synthesis in milk when whole flaxseed was administered. Short-chain fatty acids, in particular C8:0 and C10:0, were the lowest in milk from buffalo cows fed the highest level of flaxseed supplementation. Medium-chain fatty acids were the lowest in FS1000, intermediate in FS500, and the highest in the HP and LP groups. Long-chain fatty acids were the highest in FS1000, intermediate in FS500 groups, and the lowest in milk from buffalo receiving no flaxseed supplementation. Protein level of the diet influenced the percentage of C18:0, which was higher in MP than LP groups. Total conjugated linoleic acid content evidenced the same trend of long-chain fatty acids, with an increase of about 7% in FL500 and of 22% in FL1000 than the control. Apart from

  3. Quality of buffalo milk as affected by dietary protein level and flaxseed supplementation.

    PubMed

    Santillo, A; Caroprese, M; Marino, R; Sevi, A; Albenzio, M

    2016-10-01

    The aim of the present research was to evaluate the effects of protein level and flaxseed supplementation on the yield and quality of buffalo milk. In particular, the fatty acid profile of milk from buffalo cows subjected to different diets has been investigated. A 2×3 factorial design was tested with buffalo cows receiving 2 dietary crude protein (CP) and 3 flaxseed (FS) supplementation levels. Treatments were (1) low dietary CP level [12% of dry matter (DM)] and no flaxseed supplementation (LP); (2) low dietary CP level (12% of DM) and low flaxseed supplementation (500g/d) (LPFS500); (3) low dietary CP level (12% of DM) and moderate flaxseed supplementation (1,000g/d) (LPFS1000); (4) moderate dietary CP level (15% of DM) and no flaxseed supplementation (MP); (5) moderate dietary CP level (15% of DM) and low flaxseed supplementation (500g/d) (MPFS500); and (6) moderate dietary CP level (15% of DM) and moderate flaxseed supplementation (1,000g/d) (MPFS1000). Milk protein and casein were affected by flaxseed supplementation being higher in MP, intermediate in LP, and lower in flaxseed-supplemented diets. However, the results from the present study highlighted that low protein diets sustained milk yield, protein, and casein synthesis in milk when whole flaxseed was administered. Short-chain fatty acids, in particular C8:0 and C10:0, were the lowest in milk from buffalo cows fed the highest level of flaxseed supplementation. Medium-chain fatty acids were the lowest in FS1000, intermediate in FS500, and the highest in the HP and LP groups. Long-chain fatty acids were the highest in FS1000, intermediate in FS500 groups, and the lowest in milk from buffalo receiving no flaxseed supplementation. Protein level of the diet influenced the percentage of C18:0, which was higher in MP than LP groups. Total conjugated linoleic acid content evidenced the same trend of long-chain fatty acids, with an increase of about 7% in FL500 and of 22% in FL1000 than the control. Apart from

  4. Understanding of how Propionibacterium acidipropionici respond to propionic acid stress at the level of proteomics

    PubMed Central

    Guan, Ningzi; Shin, Hyun-dong; Chen, Rachel R.; Li, Jianghua; Liu, Long; Du, Guocheng; Chen, Jian

    2014-01-01

    Propionic acid (PA) is an important platform chemical in the food, agriculture, and pharmaceutical industries and is mainly biosynthesized by propionibacteria. Acid tolerance in PA-producing strains is crucial. In previous work, we investigated the acid tolerance mechanism of Propionibacterium acidipropionici at microenvironmental levels by analyzing physiological changes in the parental strain and three PA-tolerant mutants obtained by genome shuffling. However, the molecular mechanism of PA tolerance in P. acidipropionici remained unclear. Here, we performed a comparative proteomics study of P. acidipropionici CGMCC 1.2230 and the acid-tolerant mutant P. acidipropionici WSH1105; MALDI-TOF/MS identified 24 proteins that significantly differed between the parental and shuffled strains. The differentially expressed proteins were mainly categorized as key components of crucial biological processes and the acid stress response. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to confirm differential expression of nine key proteins. Overexpression of the secretory protein glyceraldehyde-3-phosphate dehydrogenase and ATP synthase subunit α in Escherichia coli BL21 improved PA and acetic acid tolerance; overexpression of NADH dehydrogenase and methylmalonyl-CoA epimerase improved PA tolerance. These results provide new insights into the acid tolerance of P. acidipropionici and will facilitate the development of PA production through fermentation by propionibacteria. PMID:25377721

  5. Dietary protein source and level alters growth in neon tetras.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nutritional studies for aquarium fish like the neon tetra are sparse in comparison with those for food fish. To determine the optimum dietary protein level and source for growth of neon tetras, diets were formulated to contain 25, 35, 45 and 55% dietary protein from either marine animal protein or ...

  6. Olive oils modulate fatty acid content and signaling protein expression in apolipoprotein E knockout mice brain.

    PubMed

    Alemany, Regina; Navarro, María A; Vögler, Oliver; Perona, Javier S; Osada, Jesús; Ruiz-Gutiérrez, Valentina

    2010-01-01

    Atherosclerosis contributes to disruption of neuronal signaling pathways by producing lipid-dependent modifications of brain plasma membranes, neuroinflammation and oxidative stress. We investigated whether long-term (11 weeks) consumption of refined- (ROO) and pomace- (POO) olive oil modulated the fatty acid composition and the levels of membrane signaling proteins in the brain of apolipoprotein E (apoE) knockout (KO) mice, an animal model of atherosclerosis. Both of these oils are rich in bioactive molecules with anti-inflammatory and antioxidant effects. ROO and POO long-term consumption increased the proportion of monounsaturated fatty acids (MUFAs), particularly of oleic acid, while reducing the level of the saturated fatty acids (SFAs) palmitic and stearic acid. As a result, the MUFA:SFA ratio was higher in apoE KO mice brain fed with ROO and POO. Furthermore, both oils reduced the level of arachidonic and eicosapentaenoic acid, suggesting a decrease in the generation of pro- and anti-inflammatory eicosanoids. Finally, ROO and POO induced an increase in the density of membrane proteins implicated in both the Galphas/PKA and Galphaq/PLCbeta1/PKCalpha signaling pathways. The combined effects of long-term ROO and POO consumption on fatty acid composition and the level of signaling proteins involved in PKA and PKC activation, suggest positive effects on neuroinflammation and brain function in apoE KO mice brain, and convert these oils into promising functional foods in diseases involving apoE deficiency.

  7. [Hydroxycinnamic acid levels of various batches from mugwort flowering tops].

    PubMed

    Fraisse, D; Carnat, A; Carnat, A-P; Guédon, D; Lamaison, J-L

    2003-07-01

    Dried flowering tops of 24 harvested batches (Artemisia vulgaris: 13; Artemisia verlotiorum: 11) and 12 batches of mugwort from commercial origin were examined. The levels of principal compounds averaged respectively: total hydroxycinnamic acids 6.09; 10.29 and 9.13%, chlorogenic acid 0.79; 2.05 and 1.35%, 1,5-dicaffeoylquinic acid 0.51; 4.01 and 1.25%, 3,5-dicaffeoylquinic acid 2.21; 1.25 and 2.60%. Specifications were discussed for an European Pharmacopoeial monography.

  8. [Hydroxycinnamic acid levels of various batches from mugwort flowering tops].

    PubMed

    Fraisse, D; Carnat, A; Carnat, A-P; Guédon, D; Lamaison, J-L

    2003-07-01

    Dried flowering tops of 24 harvested batches (Artemisia vulgaris: 13; Artemisia verlotiorum: 11) and 12 batches of mugwort from commercial origin were examined. The levels of principal compounds averaged respectively: total hydroxycinnamic acids 6.09; 10.29 and 9.13%, chlorogenic acid 0.79; 2.05 and 1.35%, 1,5-dicaffeoylquinic acid 0.51; 4.01 and 1.25%, 3,5-dicaffeoylquinic acid 2.21; 1.25 and 2.60%. Specifications were discussed for an European Pharmacopoeial monography. PMID:12843960

  9. Predicting protein disorder by analyzing amino acid sequence

    PubMed Central

    Yang, Jack Y; Yang, Mary Qu

    2008-01-01

    Background Many protein regions and some entire proteins have no definite tertiary structure, presenting instead as dynamic, disorder ensembles under different physiochemical circumstances. These proteins and regions are known as Intrinsically Unstructured Proteins (IUP). IUP have been associated with a wide range of protein functions, along with roles in diseases characterized by protein misfolding and aggregation. Results Identifying IUP is important task in structural and functional genomics. We exact useful features from sequences and develop machine learning algorithms for the above task. We compare our IUP predictor with PONDRs (mainly neural-network-based predictors), disEMBL (also based on neural networks) and Globplot (based on disorder propensity). Conclusion We find that augmenting features derived from physiochemical properties of amino acids (such as hydrophobicity, complexity etc.) and using ensemble method proved beneficial. The IUP predictor is a viable alternative software tool for identifying IUP protein regions and proteins. PMID:18831799

  10. IR-UV photochemistry of protein-nucleic acid systems

    SciTech Connect

    Kozub, J.; Edwards, G.

    1995-12-31

    UV light has often been used to induce the formation of covalent bonds between DNA (or RNA) and tightly-bound protein molecules. However, the internal photoreactions of nucleic acids and proteins limit the yield and complicate the analysis of intermolecular crosslinks. In an ongoing search for improved reaction specificity or new photoreactions in these systems, we have employed UV photons from a Nd:YAG-pumped dye laser and mid-IR photons from the Vanderbilt FEL. Having crosslinked several protein-nucleic acid systems with nanosecond UV laser pulses, we are currently studying the effect of various IR wavelengths on a model system (gene 32 protein and poly[dT]). We have found that irradiation with sufficiently intense FEL macropulses creates an altered form of gene 32 protein which was not observed with UV-only irradiation. The electrophoretic nobility of the product is consistent with the formation of a specific protein-protein crosslink. No evidence of the non-specific protein damage typically induced by UV light is found. The yield of the new photoproduct is apparently enhanced by exposure to FEL macropulses which are synchronized with UV laser pulses. With ideal exposure parameters, the two-color reaction effectively competes with UV-only reactions. Experiments designed to determine the reaction mechanism and to demonstrate FEL-induced reactions in other protein-nucleic acid systems are currently underway.

  11. Molecular Evolution Directs Protein Translation Using Unnatural Amino Acids.

    PubMed

    Cox, Vanessa E; Gaucher, Eric A

    2015-12-02

    Unnatural amino acids have in recent years established their importance in a wide range of fields, from pharmaceuticals to polymer science. Unnatural amino acids can increase the number of chemical groups within proteins and thus expand or enhance biological function. Our ability to utilize these important building blocks, however, has been limited by the inherent difficulty in incorporating these molecules into proteins. To address this challenge, researchers have examined how the canonical twenty amino acids are incorporated, regulated, and modified in nature. This review focuses on achievements and techniques used to engineer the ribosomal protein-translation machinery, including the introduction of orthogonal translation components, how directed evolution enhances the incorporation of unnatural amino acids, and the potential utility of ancient biomolecules for this process.

  12. Modifications of proteins by polyunsaturated fatty acid peroxidation products

    NASA Astrophysics Data System (ADS)

    Refsgaard, Hanne H. F.; Tsai, Lin; Stadtman, Earl R.

    2000-01-01

    The ability of unsaturated fatty acid methyl esters to modify amino acid residues in bovine serum albumin (BSA), glutamine synthetase, and insulin in the presence of a metal-catalyzed oxidation system [ascorbate/Fe(III)/O2] depends on the degree of unsaturation of the fatty acid. The fatty acid-dependent generation of carbonyl groups and loss of lysine residues increased in the order methyl linoleate < methyl linolenate < methyl arachidonate. The amounts of alkyl hydroperoxides, malondialdehyde, and a number of other aldehydes that accumulated when polyunsaturated fatty acids were oxidized in the presence of BSA were significantly lower than that observed in the absence of BSA. Direct treatment of proteins with various lipid hydroperoxides led to a slight increase in the formation of protein carbonyl derivatives, whereas treatment with the hydroperoxides together with Fe(II) led to a substantial increase in the formation of protein carbonyls. These results are consistent with the proposition that metal-catalyzed oxidation of polyunsaturated fatty acids can contribute to the generation of protein carbonyls by direct interaction of lipid oxidation products (α,β-unsaturated aldehydes) with lysine residues (Michael addition reactions) and also by interactions with alkoxyl radicals obtained by Fe(II) cleavage of lipid hydroperoxides that are formed. In addition, saturated aldehydes derived from the polyunsaturated fatty acids likely react with lysine residues to form Schiff base adducts.

  13. Modifications of proteins by polyunsaturated fatty acid peroxidation products

    PubMed Central

    Refsgaard, Hanne H. F.; Tsai, Lin; Stadtman, Earl R.

    2000-01-01

    The ability of unsaturated fatty acid methyl esters to modify amino acid residues in bovine serum albumin (BSA), glutamine synthetase, and insulin in the presence of a metal-catalyzed oxidation system [ascorbate/Fe(III)/O2] depends on the degree of unsaturation of the fatty acid. The fatty acid-dependent generation of carbonyl groups and loss of lysine residues increased in the order methyl linoleate < methyl linolenate < methyl arachidonate. The amounts of alkyl hydroperoxides, malondialdehyde, and a number of other aldehydes that accumulated when polyunsaturated fatty acids were oxidized in the presence of BSA were significantly lower than that observed in the absence of BSA. Direct treatment of proteins with various lipid hydroperoxides led to a slight increase in the formation of protein carbonyl derivatives, whereas treatment with the hydroperoxides together with Fe(II) led to a substantial increase in the formation of protein carbonyls. These results are consistent with the proposition that metal-catalyzed oxidation of polyunsaturated fatty acids can contribute to the generation of protein carbonyls by direct interaction of lipid oxidation products (α,β-unsaturated aldehydes) with lysine residues (Michael addition reactions) and also by interactions with alkoxyl radicals obtained by Fe(II) cleavage of lipid hydroperoxides that are formed. In addition, saturated aldehydes derived from the polyunsaturated fatty acids likely react with lysine residues to form Schiff base adducts. PMID:10639127

  14. A nine-country study of the protein content and amino acid composition of mature human milk

    PubMed Central

    Feng, Ping; Gao, Ming; Burgher, Anita; Zhou, Tian Hui; Pramuk, Kathryn

    2016-01-01

    Background Numerous studies have evaluated protein and amino acid levels in human milk. However, research in this area has been limited by small sample sizes and study populations with little ethnic or racial diversity. Objective Evaluate the protein and amino acid composition of mature (≥30 days) human milk samples collected from a large, multinational study using highly standardized methods for sample collection, storage, and analysis. Design Using a single, centralized laboratory, human milk samples from 220 women (30–188 days postpartum) from nine countries were analyzed for amino acid composition using Waters AccQ-Tag high-performance liquid chromatography and total nitrogen content using the LECO FP-528 nitrogen analyzer. Total protein was calculated as total nitrogen×6.25. True protein, which includes protein, free amino acids, and peptides, was calculated from the total amino acids. Results Mean total protein from individual countries (standard deviation [SD]) ranged from 1,133 (125.5) to 1,366 (341.4) mg/dL; the mean across all countries (SD) was 1,192 (200.9) mg/dL. Total protein, true protein, and amino acid composition were not significantly different across countries except Chile, which had higher total and true protein. Amino acid profiles (percent of total amino acids) did not differ across countries. Total and true protein concentrations and 16 of 18 amino acid concentrations declined with the stage of lactation. Conclusions Total protein, true protein, and individual amino acid concentrations in human milk steadily decline from 30 to 151 days of lactation, and are significantly higher in the second month of lactation compared with the following 4 months. There is a high level of consistency in the protein content and amino acid composition of human milk across geographic locations. The size and diversity of the study population and highly standardized procedures for the collection, storage, and analysis of human milk support the validity and

  15. Chemical approaches to detect and analyze protein sulfenic acids

    PubMed Central

    Furdui, Cristina M.; Poole, Leslie B.

    2013-01-01

    Orchestration of many processes relying on intracellular signal transduction is recognized to require the generation of hydrogen peroxide as a second messenger, yet relatively few molecular details of how this oxidant acts to regulate protein function are currently understood. This review describes emerging chemical tools and approaches that can be applied to study protein oxidation in biological systems, with a particular emphasis on a key player in protein redox regulation, cysteine sulfenic acid. While sulfenic acids (within purified proteins or simple mixtures) are detectable by physical approaches like X-ray crystallography, nuclear magnetic resonance and mass spectrometry, the propensity of these moieties to undergo further modification in complex biological systems has necessitated the development of chemical probes, reporter groups and analytical approaches to allow for their selective detection and quantification. Provided is an overview of techniques that are currently available for the study of sulfenic acids, and some of the biologically meaningful data that have been collected using such approaches. PMID:24105931

  16. Chemical approaches to detect and analyze protein sulfenic acids.

    PubMed

    Furdui, Cristina M; Poole, Leslie B

    2014-01-01

    Orchestration of many processes relying on intracellular signal transduction is recognized to require the generation of hydrogen peroxide as a second messenger, yet relatively few molecular details of how this oxidant acts to regulate protein function are currently understood. This review describes emerging chemical tools and approaches that can be applied to study protein oxidation in biological systems, with a particular emphasis on a key player in protein redox regulation, cysteine sulfenic acid. While sulfenic acids (within purified proteins or simple mixtures) are detectable by physical approaches like X-ray crystallography, nuclear magnetic resonance and mass spectrometry, the propensity of these moieties to undergo further modification in complex biological systems has necessitated the development of chemical probes, reporter groups and analytical approaches to allow for their selective detection and quantification. Provided is an overview of techniques that are currently available for the study of sulfenic acids, and some of the biologically meaningful data that have been collected using such approaches.

  17. On combining protein sequences and nucleic acid sequences in phylogenetic analysis: the homeobox protein case.

    PubMed

    Agosti, D; Jacobs, D; DeSalle, R

    1996-01-01

    Amino acid encoding genes contain character state information that may be useful for phylogenetic analysis on at least two levels. The nucleotide sequence and the translated amino acid sequences have both been employed separately as character states for cladistic studies of various taxa, including studies of the genealogy of genes in multigene families. In essence, amino acid sequences and nucleic acid sequences are two different ways of character coding the information in a gene. Silent positions in the nucleotide sequence (first or third positions in codons that can accrue change without changing the identity of the amino acid that the triplet codes for) may accrue change relatively rapidly and become saturated, losing the pattern of historical divergence. On the other hand, non-silent nucleotide alterations and their accompanying amino acid changes may evolve too slowly to reveal relationships among closely related taxa. In general, the dynamics of sequence change in silent and non-silent positions in protein coding genes result in homoplasy and lack of resolution, respectively. We suggest that the combination of nucleic acid and the translated amino acid coded character states into the same data matrix for phylogenetic analysis addresses some of the problems caused by the rapid change of silent nucleotide positions and overall slow rate of change of non-silent nucleotide positions and slowly changing amino acid positions. One major theoretical problem with this approach is the apparent non-independence of the two sources of characters. However, there are at least three possible outcomes when comparing protein coding nucleic acid sequences with their translated amino acids in a phylogenetic context on a codon by codon basis. First, the two character sets for a codon may be entirely congruent with respect to the information they convey about the relationships of a certain set of taxa. Second, one character set may display no information concerning a phylogenetic

  18. Update of PROFEAT: a web server for computing structural and physicochemical features of proteins and peptides from amino acid sequence.

    PubMed

    Rao, H B; Zhu, F; Yang, G B; Li, Z R; Chen, Y Z

    2011-07-01

    Sequence-derived structural and physicochemical features have been extensively used for analyzing and predicting structural, functional, expression and interaction profiles of proteins and peptides. PROFEAT has been developed as a web server for computing commonly used features of proteins and peptides from amino acid sequence. To facilitate more extensive studies of protein and peptides, numerous improvements and updates have been made to PROFEAT. We added new functions for computing descriptors of protein-protein and protein-small molecule interactions, segment descriptors for local properties of protein sequences, topological descriptors for peptide sequences and small molecule structures. We also added new feature groups for proteins and peptides (pseudo-amino acid composition, amphiphilic pseudo-amino acid composition, total amino acid properties and atomic-level topological descriptors) as well as for small molecules (atomic-level topological descriptors). Overall, PROFEAT computes 11 feature groups of descriptors for proteins and peptides, and a feature group of more than 400 descriptors for small molecules plus the derived features for protein-protein and protein-small molecule interactions. Our computational algorithms have been extensively tested and used in a number of published works for predicting proteins of specific structural or functional classes, protein-protein interactions, peptides of specific functions and quantitative structure activity relationships of small molecules. PROFEAT is accessible free of charge at http://bidd.cz3.nus.edu.sg/cgi-bin/prof/protein/profnew.cgi.

  19. Nucleic acid compositions and the encoding proteins

    SciTech Connect

    Preston, III, James F.; Chow, Virginia; Nong, Guang; Rice, John D.; St. John, Franz J.

    2014-09-02

    The subject invention provides at least one nucleic acid sequence encoding an aldouronate-utilization regulon isolated from Paenibacillus sp. strain JDR-2, a bacterium which efficiently utilizes xylan and metabolizes aldouronates (methylglucuronoxylosaccharides). The subject invention also provides a means for providing a coordinately regulated process in which xylan depolymerization and product assimilation are coupled in Paenibacillus sp. strain JDR-2 to provide a favorable system for the conversion of lignocellulosic biomass to biobased products. Additionally, the nucleic acid sequences encoding the aldouronate-utilization regulon can be used to transform other bacteria to form organisms capable of producing a desired product (e.g., ethanol, 1-butanol, acetoin, 2,3-butanediol, 1,3-propanediol, succinate, lactate, acetate, malate or alanine) from lignocellulosic biomass.

  20. Mapping protein and nucleic acid structure

    NASA Astrophysics Data System (ADS)

    Bednyakov, I. V.; Zrelov, P. V.; Ivanov, V. V.; Polozov, R. V.; Sivozhelezov, V. S.; Stepanenko, V. A.; Chirgadze, Yu. N.

    2013-09-01

    Methods and algorithms to analyze surfaces of globular and fibrillar proteins, DNA, and RNA have been developed. These methods for the construction of maps of fragments of these objects in the original cylindrical projection developed herein essentially broaden the possibilities for studying the distribution of charges and surface topography of biological structures. This approach significantly supplements the qualitative characteristics of methods of visualizing biopolymer structures.

  1. Cinnamic acid and its derivatives inhibit fructose-mediated protein glycation.

    PubMed

    Adisakwattana, Sirichai; Sompong, Weerachat; Meeprom, Aramsri; Ngamukote, Sathaporn; Yibchok-Anun, Sirintorn

    2012-01-01

    Cinnamic acid and its derivatives have shown a variety of pharmacologic properties. However, little is known about the antiglycation properties of cinnamic acid and its derivatives. The present study sought to characterize the protein glycation inhibitory activity of cinnamic acid and its derivatives in a bovine serum albumin (BSA)/fructose system. The results demonstrated that cinnamic acid and its derivatives significantly inhibited the formation of advanced glycation end products (AGEs) by approximately 11.96-63.36% at a concentration of 1 mM. The strongest inhibitory activity against the formation of AGEs was shown by cinnamic acid. Furthermore, cinnamic acid and its derivatives reduced the level of fructosamine, the formation of N(ɛ)-(carboxymethyl) lysine (CML), and the level of amyloid cross β-structure. Cinnamic acid and its derivatives also prevented oxidative protein damages, including effects on protein carbonyl formation and thiol oxidation of BSA. Our findings may lead to the possibility of using cinnamic acid and its derivatives for preventing AGE-mediated diabetic complications.

  2. Abscisic acid (ABA) regulation of Arabidopsis SR protein gene expression.

    PubMed

    Cruz, Tiago M D; Carvalho, Raquel F; Richardson, Dale N; Duque, Paula

    2014-01-01

    Serine/arginine-rich (SR) proteins are major modulators of alternative splicing, a key generator of proteomic diversity and flexible means of regulating gene expression likely to be crucial in plant environmental responses. Indeed, mounting evidence implicates splicing factors in signal transduction of the abscisic acid (ABA) phytohormone, which plays pivotal roles in the response to various abiotic stresses. Using real-time RT-qPCR, we analyzed total steady-state transcript levels of the 18 SR and two SR-like genes from Arabidopsis thaliana in seedlings treated with ABA and in genetic backgrounds with altered expression of the ABA-biosynthesis ABA2 and the ABA-signaling ABI1 and ABI4 genes. We also searched for ABA-responsive cis elements in the upstream regions of the 20 genes. We found that members of the plant-specific SC35-Like (SCL) Arabidopsis SR protein subfamily are distinctively responsive to exogenous ABA, while the expression of seven SR and SR-related genes is affected by alterations in key components of the ABA pathway. Finally, despite pervasiveness of established ABA-responsive promoter elements in Arabidopsis SR and SR-like genes, their expression is likely governed by additional, yet unidentified cis-acting elements. Overall, this study pinpoints SR34, SR34b, SCL30a, SCL28, SCL33, RS40, SR45 and SR45a as promising candidates for involvement in ABA-mediated stress responses. PMID:25268622

  3. Abscisic Acid (ABA) Regulation of Arabidopsis SR Protein Gene Expression

    PubMed Central

    Cruz, Tiago M. D.; Carvalho, Raquel F.; Richardson, Dale N.; Duque, Paula

    2014-01-01

    Serine/arginine-rich (SR) proteins are major modulators of alternative splicing, a key generator of proteomic diversity and flexible means of regulating gene expression likely to be crucial in plant environmental responses. Indeed, mounting evidence implicates splicing factors in signal transduction of the abscisic acid (ABA) phytohormone, which plays pivotal roles in the response to various abiotic stresses. Using real-time RT-qPCR, we analyzed total steady-state transcript levels of the 18 SR and two SR-like genes from Arabidopsis thaliana in seedlings treated with ABA and in genetic backgrounds with altered expression of the ABA-biosynthesis ABA2 and the ABA-signaling ABI1 and ABI4 genes. We also searched for ABA-responsive cis elements in the upstream regions of the 20 genes. We found that members of the plant-specific SC35-Like (SCL) Arabidopsis SR protein subfamily are distinctively responsive to exogenous ABA, while the expression of seven SR and SR-related genes is affected by alterations in key components of the ABA pathway. Finally, despite pervasiveness of established ABA-responsive promoter elements in Arabidopsis SR and SR-like genes, their expression is likely governed by additional, yet unidentified cis-acting elements. Overall, this study pinpoints SR34, SR34b, SCL30a, SCL28, SCL33, RS40, SR45 and SR45a as promising candidates for involvement in ABA-mediated stress responses. PMID:25268622

  4. The Hsp90-Dependent Proteome Is Conserved and Enriched for Hub Proteins with High Levels of Protein–Protein Connectivity

    PubMed Central

    Swamy, Krishna B.S.; Yu, Jau-Song; Schuyler, Scott C.; Leu, Jun-Yi

    2014-01-01

    Hsp90 is one of the most abundant and conserved proteins in the cell. Reduced levels or activity of Hsp90 causes defects in many cellular processes and also reveals genetic and nongenetic variation within a population. Despite information about Hsp90 protein–protein interactions, a global view of the Hsp90-regulated proteome in yeast is unavailable. To investigate the degree of dependency of individual yeast proteins on Hsp90, we used the “stable isotope labeling by amino acids in cell culture” method coupled with mass spectrometry to quantify around 4,000 proteins in low-Hsp90 cells. We observed that 904 proteins changed in their abundance by more than 1.5-fold. When compared with the transcriptome of the same population of cells, two-thirds of the misregulated proteins were observed to be affected posttranscriptionally, of which the majority were downregulated. Further analyses indicated that the downregulated proteins are highly conserved and assume central roles in cellular networks with a high number of protein interacting partners, suggesting that Hsp90 buffers genetic and nongenetic variation through regulating protein network hubs. The downregulated proteins were enriched for essential proteins previously not known to be Hsp90-dependent. Finally, we observed that downregulation of transcription factors and mating pathway components by attenuating Hsp90 function led to decreased target gene expression and pheromone response, respectively, providing a direct link between observed proteome regulation and cellular phenotypes. PMID:25316598

  5. FLU, an amino acid substitution model for influenza proteins

    PubMed Central

    2010-01-01

    Background The amino acid substitution model is the core component of many protein analysis systems such as sequence similarity search, sequence alignment, and phylogenetic inference. Although several general amino acid substitution models have been estimated from large and diverse protein databases, they remain inappropriate for analyzing specific species, e.g., viruses. Emerging epidemics of influenza viruses raise the need for comprehensive studies of these dangerous viruses. We propose an influenza-specific amino acid substitution model to enhance the understanding of the evolution of influenza viruses. Results A maximum likelihood approach was applied to estimate an amino acid substitution model (FLU) from ~113, 000 influenza protein sequences, consisting of ~20 million residues. FLU outperforms 14 widely used models in constructing maximum likelihood phylogenetic trees for the majority of influenza protein alignments. On average, FLU gains ~42 log likelihood points with an alignment of 300 sites. Moreover, topologies of trees constructed using FLU and other models are frequently different. FLU does indeed have an impact on likelihood improvement as well as tree topologies. It was implemented in PhyML and can be downloaded from ftp://ftp.sanger.ac.uk/pub/1000genomes/lsq/FLU or included in PhyML 3.0 server at http://www.atgc-montpellier.fr/phyml/. Conclusions FLU should be useful for any influenza protein analysis system which requires an accurate description of amino acid substitutions. PMID:20384985

  6. Effect of Dietary Protein Levels on Composition of Odorous Compounds and Bacterial Ecology in Pig Manure.

    PubMed

    Cho, Sungback; Hwang, Okhwa; Park, Sungkwon

    2015-09-01

    This study was performed to investigate the effect of different levels of dietary crude protein (CP) on composition of odorous compounds and bacterial communities in pig manure. A total of 48 male pigs (average initial body weight 45 kg) fed diets containing three levels of dietary CP (20%, 17.5%, and 15%) and their slurry samples were collected from the pits under the floor every week for one month. Changes in composition of odorous compounds and bacterial communities were analyzed by gas chromatography and 454 FLX titanium pyrosequencing systems, respectively. Levels of phenols, indoles, short chain fatty acid and branched chain fatty acid were lowest (p<0.05) in CP 15% group among three CP levels. Relative abundance of Bacteroidetes phylum and bacterial genera including Leuconostoc, Bacillus, Atopostipes, Peptonphilus, Ruminococcaceae_uc, Bacteroides, and Pseudomonas was lower (p<0.05) in CP 15% than in CP 20% group. There was a positive correlation (p<0.05) between odorous compounds and bacterial genera: phenol, indole, iso-butyric acid, and iso-valeric acid with Atopostipes, p-cresol and skatole with Bacteroides, acetic acid and butyric acid with AM982595_g of Porphyromonadaceae family, and propionic acid with Tissierella. Taken together, administration of 15% CP showed less production of odorous compounds than 20% CP group and this result might be associated with the changes in bacterial communities especially whose roles in protein metabolism. PMID:26194219

  7. Effect of Dietary Protein Levels on Composition of Odorous Compounds and Bacterial Ecology in Pig Manure

    PubMed Central

    Cho, Sungback; Hwang, Okhwa; Park, Sungkwon

    2015-01-01

    This study was performed to investigate the effect of different levels of dietary crude protein (CP) on composition of odorous compounds and bacterial communities in pig manure. A total of 48 male pigs (average initial body weight 45 kg) fed diets containing three levels of dietary CP (20%, 17.5%, and 15%) and their slurry samples were collected from the pits under the floor every week for one month. Changes in composition of odorous compounds and bacterial communities were analyzed by gas chromatography and 454 FLX titanium pyrosequencing systems, respectively. Levels of phenols, indoles, short chain fatty acid and branched chain fatty acid were lowest (p<0.05) in CP 15% group among three CP levels. Relative abundance of Bacteroidetes phylum and bacterial genera including Leuconostoc, Bacillus, Atopostipes, Peptonphilus, Ruminococcaceae_uc, Bacteroides, and Pseudomonas was lower (p<0.05) in CP 15% than in CP 20% group. There was a positive correlation (p<0.05) between odorous compounds and bacterial genera: phenol, indole, iso-butyric acid, and iso-valeric acid with Atopostipes, p-cresol and skatole with Bacteroides, acetic acid and butyric acid with AM982595_g of Porphyromonadaceae family, and propionic acid with Tissierella. Taken together, administration of 15% CP showed less production of odorous compounds than 20% CP group and this result might be associated with the changes in bacterial communities especially whose roles in protein metabolism. PMID:26194219

  8. Stability of free amino acid levels in stressed Abarenicola pacifica

    SciTech Connect

    Augenfeld, J.M.; Anderson, J.W.

    1980-01-01

    A change in the composition of the free amino acid (FAA) pool of pelecypod molluscs under stress has been described. This response consists of a significant decrease in the level of glycine, while the taurine level remains constant, leading to a decrease in the total free amino acid level and a decrease in the glycine:taurine ratio. It has been suggested that this alteration in FAA composition may be useful as an indicator of physiological stress in a wide variety of invertebrates. This suggestion has been tested by exposing a burrowing polychaete, Abarenicola pacifica, to sediment artificially contaminated with Prudhoe Bay crude oil (PBC).

  9. Review: the liver bile acid-binding proteins.

    PubMed

    Monaco, Hugo L

    2009-12-01

    The liver bile acid-binding proteins, L-BABPs, formerly called the liver "basic" fatty acid-binding proteins, are a subfamily of the fatty acid-binding proteins, FABPs. All the members of this protein group share the same fold: a 10 stranded beta barrel in which two short helices are inserted in between the first and the second strand of antiparallel beta sheet. The barrel encloses the ligand binding cavity of the protein while the two helices are believed to be involved in ligand accessibility to the binding site. The L-BABP subfamily has been found to be present in the liver of several vertebrates: fish, amphibians, reptiles, and birds but not in mammals. The members of the FABP family present in mammals that appear to be more closely related to the L-BABPs are the liver FABPs and the ileal BABPs, both very extensively studied. Several L-BABP X-ray structures are available and chicken L-BABP has also been studied using NMR spectroscopy. The stoichiometry of ligand binding for bile acids, first determined by X-ray crystallography for the chicken liver protein, is of two cholates per protein molecule with the only exception of zebrafish L-BABP which, due to the presence of a disulfide bridge, has a stoichiometry of 1:1. The stoichiometry of ligand binding for fatty acids, determined with several different techniques, is 1:1. An unanswered question of great relevance is the identity of the protein that in mammals performs the function that in other vertebrates is carried out by the L-BABPS.

  10. Gut Microbial Fatty Acid Metabolites Reduce Triacylglycerol Levels in Hepatocytes.

    PubMed

    Nanthirudjanar, Tharnath; Furumoto, Hidehiro; Zheng, Jiawen; Kim, Young-Il; Goto, Tsuyoshi; Takahashi, Nobuyuki; Kawada, Teruo; Park, Si-Bum; Hirata, Akiko; Kitamura, Nahoko; Kishino, Shigenobu; Ogawa, Jun; Hirata, Takashi; Sugawara, Tatsuya

    2015-11-01

    Hydroxy and oxo fatty acids were recently found to be produced as intermediates during gut microbial fatty acid metabolism. Lactobacillus plantarum produces these fatty acids from unsaturated fatty acids such as linoleic acid. In this study, we investigated the effects of these gut microbial fatty acid metabolites on the lipogenesis in liver cells. We screened their effect on sterol regulatory element binding protein-1c (SREBP-1c) expression in HepG2 cells treated with a synthetic liver X receptor α (LXRα) agonist (T0901317). The results showed that 10-hydroxy-12(Z)-octadecenoic acid (18:1) (HYA), 10-hydroxy-6(Z),12(Z)-octadecadienoic acid (18:2) (γHYA), 10-oxo-12(Z)-18:1 (KetoA), and 10-oxo-6(Z),12(Z)-18:2 (γKetoA) significantly decreased SREBP-1c mRNA expression induced by T0901317. These fatty acids also downregulated the mRNA expression of lipogenic genes by suppressing LXRα activity and inhibiting SREBP-1 maturation. Oral administration of KetoA, which effectively reduced triacylglycerol accumulation and acetyl-CoA carboxylase 2 (ACC2) expression in HepG2 cells, for 2 weeks significantly decreased Srebp-1c, Scd-1, and Acc2 expression in the liver of mice fed a high-sucrose diet. Our findings suggest that the hypolipidemic effect of the fatty acid metabolites produced by L. plantarum can be exploited in the treatment of cardiovascular diseases or dyslipidemia. PMID:26399511

  11. [Fractional and amino acid composition of krill proteins and the potential for obtaining protein preparations].

    PubMed

    Orlova, T A; Churina, E E; Kuranova, L K

    1985-01-01

    Studies of the fractional composition of krill proteins demonstrated that the content of protein fractions changes depending on the time of krill catch. The highest amount of water-soluble proteins is contained by krill caught in December (64%), of salt-soluble by krill caught in June (12%), base-soluble by krill caught in May, September and February (34%). Krill protein contains from 50 to 60% of water- and salt-soluble fractions. Analysis of the amino acid composition of krill proteins showed that it does not differ essentially from that of adequate food proteins.

  12. Differential regulation of protein synthesis by amino acids and insulin in peripheral and visceral tissues of neonatal pigs.

    PubMed

    Suryawan, Agus; O'Connor, Pamela M J; Bush, Jill A; Nguyen, Hanh V; Davis, Teresa A

    2009-05-01

    The high efficiency of protein deposition during the neonatal period is driven by high rates of protein synthesis, which are maximally stimulated after feeding. In the current study, we examined the individual roles of amino acids and insulin in the regulation of protein synthesis in peripheral and visceral tissues of the neonate by performing pancreatic glucose-amino acid clamps in overnight-fasted 7-day-old pigs. We infused pigs (n = 8-12/group) with insulin at 0, 10, 22, and 110 ng kg(-0.66) min(-1) to achieve approximately 0, 2, 6 and 30 muU ml(-1) insulin so as to simulate below fasting, fasting, intermediate, and fed insulin levels, respectively. At each insulin dose, amino acids were maintained at the fasting or fed level. In conjunction with the highest insulin dose, amino acids were also allowed to fall below the fasting level. Tissue protein synthesis was measured using a flooding dose of L: -[4-(3)H] phenylalanine. Both insulin and amino acids increased fractional rates of protein synthesis in longissimus dorsi, gastrocnemius, masseter, and diaphragm muscles. Insulin, but not amino acids, increased protein synthesis in the skin. Amino acids, but not insulin, increased protein synthesis in the liver, pancreas, spleen, and lung and tended to increase protein synthesis in the jejunum and kidney. Neither insulin nor amino acids altered protein synthesis in the stomach. The results suggest that the stimulation of protein synthesis by feeding in most tissues of the neonate is regulated by the post-prandial rise in amino acids. However, the feeding-induced stimulation of protein synthesis in skeletal muscles is independently mediated by insulin as well as amino acids.

  13. Proteins, Peptides and Amino Acids: Role in Infant Nutrition.

    PubMed

    Nutten, Sophie

    2016-01-01

    Proteins are polymers composed of 30 or more amino acids; some of them are essential dietary components, since they are not synthetized by human metabolic processes. They are crucial for healthy growth and development and influence major functions of the body. The infant's first year is a critical time of rapid growth and development, which must be supported by a high rate of protein synthesis. Breast milk, as a single specific food source in the first months of life, is providing the total protein and essential amino acids required. Infant formulas have been designed for infants who cannot be breastfed. They should be similar to breast milk in their composition and their functional outcomes, insuring appropriate growth, optimal development, maturation of the immune system, easy digestion and healthy metabolic programming. By modifying their protein components, specific infant formulas have also been developed for specific needs. For example, partially hydrolyzed (prevention of atopic dermatitis) and extensively hydrolyzed or amino-acid-based infant formulas (reduction in allergy symptoms) have been designed for the management of cow's milk protein allergy. In conclusion, proteins provided via breast milk or infant formula are essential components of the infant's diet; therefore, the specific quality, quantity and conformation of proteins are of utmost importance for healthy growth and development. PMID:27336588

  14. Proteins, Peptides and Amino Acids: Role in Infant Nutrition.

    PubMed

    Nutten, Sophie

    2016-01-01

    Proteins are polymers composed of 30 or more amino acids; some of them are essential dietary components, since they are not synthetized by human metabolic processes. They are crucial for healthy growth and development and influence major functions of the body. The infant's first year is a critical time of rapid growth and development, which must be supported by a high rate of protein synthesis. Breast milk, as a single specific food source in the first months of life, is providing the total protein and essential amino acids required. Infant formulas have been designed for infants who cannot be breastfed. They should be similar to breast milk in their composition and their functional outcomes, insuring appropriate growth, optimal development, maturation of the immune system, easy digestion and healthy metabolic programming. By modifying their protein components, specific infant formulas have also been developed for specific needs. For example, partially hydrolyzed (prevention of atopic dermatitis) and extensively hydrolyzed or amino-acid-based infant formulas (reduction in allergy symptoms) have been designed for the management of cow's milk protein allergy. In conclusion, proteins provided via breast milk or infant formula are essential components of the infant's diet; therefore, the specific quality, quantity and conformation of proteins are of utmost importance for healthy growth and development.

  15. Evaluation of circulating levels and renal clearance of natural amino acids in patients with Cushing's disease.

    PubMed

    Faggiano, A; Pivonello, R; Melis, D; Alfieri, R; Filippella, M; Spagnuolo, G; Salvatore, F; Lombardi, G; Colao, A

    2002-02-01

    Although the hypercortisolism-induced impairment of protein homeostasis is object of several studies, a detailed evaluation of the complete amino acid profile of patients with Cushing's syndrome (CS) has never been performed. The aim of the current open transversal controlled study was to evaluate serum and urinary concentrations as well as renal clearance of the complete series of natural amino acids and their relationship with glucose tolerance in patients with Cushing's disease (CD). Twenty patients with CD (10 active and 10 cured) and 20 sex- and age-matched healthy controls entered the study. Measurement of serum and urinary levels of the complete series of natural amino acids was performed in all patients analyzed by cationic exchange high performance liquid cromatography (HPLC) after 2 weeks of a standardized protein intake regimen. The renal clearance (renal excretion rate) of each amino acid was calculated on the basis of the serum and urinary concentrations of creatinine and the specific amino acid. Fasting glucose and insulin levels, glucose and insulin response to standard glucose load, insulinogenic and homeostasis model insulin resistance (Homa-R) indexes were also evaluated and correlated to the circulating levels and renal clearances of each amino acid. Significantly higher serum (p<0.01) and urinary (p<0.05) levels of alanine and cystine, lower serum and higher urinary levels of leucine, isoleucine and valine (p<0.05) and higher renal excretion rates of leucine, isoleucine and valine (p<0.01) were found in patients with active CD than in patients cured from the disease and in controls. No difference was found between cured patients and controls. Creatinine clearance was similar in active and cured patients and in controls. In patients with active CD, urinary cortisol levels were significantly correlated to urinary cystine levels (r=0.85; p<0.01) and renal excretion rate of leucine (r=-0.76; p<0.05), isoleucine (r=-0.76; p<0.05) and valine (r=-0

  16. A general method of protein purification for recombinant unstructured non-acidic proteins.

    PubMed

    Campos, Francisco; Guillén, Gabriel; Reyes, José L; Covarrubias, Alejandra A

    2011-11-01

    Typical late embryogenesis abundant (LEA) proteins accumulate in response to water deficit imposed by the environment or by plant developmental programs. Because of their physicochemical properties, they can be considered as hydrophilins and as a paradigm of intrinsically unstructured proteins (IUPs) in plants. To study their biophysical and biochemical characteristics large quantities of highly purified protein are required. In this work, we report a fast and simple purification method for non-acidic recombinant LEA proteins that does not need the addition of tags and that preserves their in vitro protective activity. The method is based on the enrichment of the protein of interest by boiling the bacterial protein extract, followed by a differential precipitation with trichloroacetic acid (TCA). Using this procedure we have obtained highly pure recombinant LEA proteins of groups 1, 3, and 4 and one recombinant bacterial hydrophilin. This protocol will facilitate the purification of this type of IUPs, and could be particularly useful in proteomic projects/analyses.

  17. Amino Acid Change in the Carbohydrate Response Element Binding Protein is associated with lower triglycerides and myocardial infarction incidence depending on level of adherence to the Mediterranean diet in the PREDIMED trial

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A variant (rs3812316, C771G, and Gln241His) in the MLXIPL (Max-like protein X interacting protein-like) gene encoding the carbohydrate response element binding protein has been associated with lower triglycerides. However, its association with cardiovascular diseases and gene-diet interactions modul...

  18. Amino acid limitation induces down-regulation of WNT5a at transcriptional level

    SciTech Connect

    Wang Zuguang; Chen Hong

    2009-01-23

    An aberrant WNT signaling contributes to the development and progression of multiple cancers. WNT5a is one of the WNT signaling molecules. This study was designed to test the hypothesis that amino acid deprivation induces changes in the WNT signaling pathway in colon cancer cells. Results showed that targets of the amino acid response pathway, ATF3 and p21, were induced in the human colon cancer cell line SW480 during amino acid limitation. There was a significant decrease in the WNT5a mRNA level following amino acid deprivation. The down-regulation of WNT5a mRNA by amino acid deprivation is not due to mRNA destabilization. There is a reduction of nuclear {beta}-catenin protein level by amino acid limitation. Under amino acid limitation, phosphorylation of ERK1/2 was increased and the blockage of ERK1/2 by the inhibitor U0126 partially restored WNT5a mRNA level. In conclusion, amino acid limitation in colon cancer cells induces phosphorylation of ERK1/2, which then down-regulates WNT5a expression.

  19. Molecular analysis of Xenopus laevis SPARC (Secreted Protein, Acidic, Rich in Cysteine). A highly conserved acidic calcium-binding extracellular-matrix protein.

    PubMed Central

    Damjanovski, S; Liu, F; Ringuette, M

    1992-01-01

    SPARC (Secreted Protein, Acidic, Rich in Cysteine) is expressed as a 1.6 kb mRNA in Xenopus laevis. On the basis of cDNA sequence analysis, Xenopus SPARC has a core Mr of 32643, with one potential N-glycosylation site. Western analysis of SPARC isolated from Xenopus long bone indicates that the mature protein has an Mr of 43,000. At the amino acid level, Xenopus SPARC has 78-79% sequence similarity to mouse, bovine and human SPARC. The least-conserved region is found within the N-terminal glutamic acid-rich domain, with the C-terminal Ca(2+)-binding domain being the most conserved. Adult Xenopus tissues show the same pattern of tissue-specific distribution of SPARC mRNAs as adult mouse. Images Fig. 1. Fig. 5. PMID:1736898

  20. Analysis of single nucleic acid molecules with protein nanopores

    PubMed Central

    Maglia, Giovanni; Heron, Andrew J.; Stoddart, David; Japrung, Deanpen; Bayley, Hagan

    2011-01-01

    We describe the methods used in our laboratory for the analysis of single nucleic acid molecules with protein nanopores. The technical section is preceded by a review of the variety of experiments that can be done with protein nanopores. The end goal of much of this work is single-molecule DNA sequencing, although sequencing is not discussed explicitly here. The technical section covers the equipment required for nucleic acid analysis, the preparation and storage of the necessary materials, and aspects of signal processing and data analysis. PMID:20627172

  1. Comparative proteomic analysis of differentially expressed proteins in β-aminobutyric acid enhanced Arabidopsis thaliana tolerance to simulated acid rain.

    PubMed

    Liu, Tingwu; Jiang, Xinwu; Shi, Wuliang; Chen, Juan; Pei, Zhenming; Zheng, Hailei

    2011-05-01

    Acid rain is a worldwide environmental issue that has seriously destroyed forest ecosystems. As a highly effective and broad-spectrum plant resistance-inducing agent, β-aminobutyric acid could elevate the tolerance of Arabidopsis when subjected to simulated acid rain. Using comparative proteomic strategies, we analyzed 203 significantly varied proteins of which 175 proteins were identified responding to β-aminobutyric acid in the absence and presence of simulated acid rain. They could be divided into ten groups according to their biological functions. Among them, the majority was cell rescue, development and defense-related proteins, followed by transcription, protein synthesis, folding, modification and destination-associated proteins. Our conclusion is β-aminobutyric acid can lead to a large-scale primary metabolism change and simultaneously activate antioxidant system and salicylic acid, jasmonic acid, abscisic acid signaling pathways. In addition, β-aminobutyric acid can reinforce physical barriers to defend simulated acid rain stress.

  2. Early postnatal docosahexaenoic acid levels and improved preterm brain development

    PubMed Central

    Tam, Emily W.Y.; Chau, Vann; Barkovich, A. James; Ferriero, Donna M.; Miller, Steven P.; Rogers, Elizabeth E.; Grunau, Ruth E.; Synnes, Anne R.; Xu, Duan; Foong, Justin; Brant, Rollin; Innis, Sheila M.

    2015-01-01

    Background Preterm birth has a dramatic impact on polyunsaturated fatty acid exposures for the developing brain. This study examined the association between postnatal fatty acid levels and measures of brain injury and development, as well as outcomes. Methods A cohort of 60 preterm newborns (24–32 weeks GA) was assessed using early and near-term MRI studies. Red blood cell fatty acid composition was analyzed coordinated with each scan. Outcome at a mean of 33 months corrected age was assessed using the Bayley Scales of Infant Development, 3rd edition. Results Adjusting for confounders, a 1% increase in postnatal docosahexaenoic acid (DHA) levels at early MRI was associated with 4.3-fold decreased odds of intraventricular hemorrhage, but was not associated with white matter injury or cerebellar haemorrhage. Higher DHA and lower linoleic acid (LA) levels at early MRI were associated with lower diffusivity in white matter tracts, and corresponding improved developmental scores in follow-up. Conclusion Higher DHA and lower LA levels in the first few weeks of life are associated with decreased IVH, improved microstructural brain development, and improved outcomes in preterm born children. Early, and possibly antenatal, intervention in high-risk pregnancies needs to be studied for potential benefits in preterm developmental outcomes. PMID:26761122

  3. Suppression of muscle protein turnover and amino acid degradation by dietary protein deficiency

    NASA Technical Reports Server (NTRS)

    Tawa, N. E. Jr; Goldberg, A. L.

    1992-01-01

    To define the adaptations that conserve amino acids and muscle protein when dietary protein intake is inadequate, rats (60-70 g final wt) were fed a normal or protein-deficient (PD) diet (18 or 1% lactalbumin), and their muscles were studied in vitro. After 7 days on the PD diet, both protein degradation and synthesis fell 30-40% in skeletal muscles and atria. This fall in proteolysis did not result from reduced amino acid supply to the muscle and preceded any clear decrease in plasma amino acids. Oxidation of branched-chain amino acids, glutamine and alanine synthesis, and uptake of alpha-aminoisobutyrate also fell by 30-50% in muscles and adipose tissue of PD rats. After 1 day on the PD diet, muscle protein synthesis and amino acid uptake decreased by 25-40%, and after 3 days proteolysis and leucine oxidation fell 30-45%. Upon refeeding with the normal diet, protein synthesis also rose more rapidly (+30% by 1 day) than proteolysis, which increased significantly after 3 days (+60%). These different time courses suggest distinct endocrine signals for these responses. The high rate of protein synthesis and low rate of proteolysis during the first 3 days of refeeding a normal diet to PD rats contributes to the rapid weight gain ("catch-up growth") of such animals.

  4. Structural and functional interaction of fatty acids with human liver fatty acid-binding protein (L-FABP) T94A variant.

    PubMed

    Huang, Huan; McIntosh, Avery L; Martin, Gregory G; Landrock, Kerstin K; Landrock, Danilo; Gupta, Shipra; Atshaves, Barbara P; Kier, Ann B; Schroeder, Friedhelm

    2014-05-01

    The human liver fatty acid-binding protein (L-FABP) T94A variant, the most common in the FABP family, has been associated with elevated liver triglyceride levels. How this amino acid substitution elicits these effects is not known. This issue was addressed using human recombinant wild-type (WT) and T94A variant L-FABP proteins as well as cultured primary human hepatocytes expressing the respective proteins (genotyped as TT, TC and CC). The T94A substitution did not alter or only slightly altered L-FABP binding affinities for saturated, monounsaturated or polyunsaturated long chain fatty acids, nor did it change the affinity for intermediates of triglyceride synthesis. Nevertheless, the T94A substitution markedly altered the secondary structural response of L-FABP induced by binding long chain fatty acids or intermediates of triglyceride synthesis. Finally, the T94A substitution markedly decreased the levels of induction of peroxisome proliferator-activated receptor α-regulated proteins such as L-FABP, fatty acid transport protein 5 and peroxisome proliferator-activated receptor α itself meditated by the polyunsaturated fatty acids eicosapentaenoic acid and docosahexaenoic acid in cultured primary human hepatocytes. Thus, although the T94A substitution did not alter the affinity of human L-FABP for long chain fatty acids, it significantly altered human L-FABP structure and stability, as well as the conformational and functional response to these ligands.

  5. Studies on the protein and sulfur amino acid requirements of young bobwhite quail

    USGS Publications Warehouse

    Serafin, J.A.

    1977-01-01

    Four experiments were conducted with purified diets to examine the influence of protein level and to estimate the sulfur amino acid (S.A.A.) requirement of young Bobwhite quail (Colinus virginianus). These studies demonstrated (I) that 26% protein was sufficient for rapid growth when the diet was supplemented with methionine; (2) that diets containing higher levels of protein (29.3% and 31.3%) failed to support satisfactory growth unless they contained supplemental methionine; and (3) that young Bobwhite quail require no more than 1.0% sulfur-containing amino acids for optimal growth and efficiency of feed utilization. A fifth experiment was conducted to examine the protein and S.A.A. requirements of young Bobwhite quail using practical rations and to compare results with those obtained with purified diets. Diets containing 24%, 26% and 28% protein were supplied with and without supplemental methionine in a five week study. Results showed significant growth responses to protein and supplemental methionine. Responses showed that Bobwhite quail require no more than 26% protein for maximum growth and efficiency of feed utilization when the S.A.A. level of the diet was approximately 1.0%. The results were in close agreement with those obtained with purified diets. These findings define more precisely than had been known the quantitative requirements of young Bobwhite quail for protein and for the S.A.A. necessary for optimal growth.

  6. The interaction of amino acids, peptides, and proteins with DNA.

    PubMed

    Solovyev, Andrey Y; Tarnovskaya, Svetlana I; Chernova, Irina A; Shataeva, Larisa K; Skorik, Yury A

    2015-01-01

    Amino acids that carry charges on their side groups can bind to double stranded DNA (dsDNA) and change the strength of the double helix. Measurement of the DNA melting temperature (Tm) confirmed that acidic amino acids (Glu, Asp) weaken the H-bonds between DNA strands, whereas basic amino acids (Arg, Lys) strengthen the interaction between the strands. A rank correlation exists between the amino acid isoelectric points and the observed changes in Tm. A similar dependence of the hyperchromic effect on the isoelectric point of a protein (pepsin, insulin, cortexin, and protamine) was observed for DNA-protein complexes at room temperature. Short peptides (KE, AEDG, and KEDP) containing a mixture of acidic and basic amino acid residues also affect Tm and the stability of the double helix. A model for binding Glu and Lys to dsDNA was explored by a docking simulation. The model shows that Glu, in an untwisted shape, binds to dsDNA in its major groove and disrupts three H-bonds between the strands, thereby destabilizing the double helix. Lys, in an untwisted shape, binds to the external side of the dsDNA and forms two bonds with O atoms of neighboring phosphodiester groups, thereby strengthening the DNA helix.

  7. Characterization and amino acid sequence of a fatty acid-binding protein from human heart.

    PubMed Central

    Offner, G D; Brecher, P; Sawlivich, W B; Costello, C E; Troxler, R F

    1988-01-01

    The complete amino acid sequence of a fatty acid-binding protein from human heart was determined by automated Edman degradation of CNBr, BNPS-skatole [3'-bromo-3-methyl-2-(2-nitrobenzenesulphenyl)indolenine], hydroxylamine, Staphylococcus aureus V8 proteinase, tryptic and chymotryptic peptides, and by digestion of the protein with carboxypeptidase A. The sequence of the blocked N-terminal tryptic peptide from citraconylated protein was determined by collisionally induced decomposition mass spectrometry. The protein contains 132 amino acid residues, is enriched with respect to threonine and lysine, lacks cysteine, has an acetylated valine residue at the N-terminus, and has an Mr of 14768 and an isoelectric point of 5.25. This protein contains two short internal repeated sequences from residues 48-54 and from residues 114-119 located within regions of predicted beta-structure and decreasing hydrophobicity. These short repeats are contained within two longer repeated regions from residues 48-60 and residues 114-125, which display 62% sequence similarity. These regions could accommodate the charged and uncharged moieties of long-chain fatty acids and may represent fatty acid-binding domains consistent with the finding that human heart fatty acid-binding protein binds 2 mol of oleate or palmitate/mol of protein. Detailed evidence for the amino acid sequences of the peptides has been deposited as Supplementary Publication SUP 50143 (23 pages) at the British Library Lending Division, Boston Spa, Yorkshire LS23 7BQ, U.K., from whom copies may be obtained as indicated in Biochem. J. (1988) 249, 5. PMID:3421901

  8. Influence of educational level on determinants of folic acid use.

    PubMed

    van der Pal-de Bruin, Karin M; de Walle, Hermien E K; de Rover, Carolien M; Jeeninga, Wendy; Cornel, Martina C; de Jong-van den Berg, Lolkje T W; Buitendijk, Simone E; Paulussen, Theo G W M

    2003-07-01

    In The Netherlands, periconceptional folic acid use to prevent neural tube defects was promoted through a national 'Folic Acid Campaign'. In two regions, a local campaign supplemented the national campaign to increase the chances of reaching women with low socio-economic status (SES). A framework of outcome criteria, defined as awareness knowledge, perceived safety, attitudes and subjective norms, was developed to evaluate the effectiveness of the two local campaigns. Data were gathered by means of two cross-sectional studies conducted just before and 1 year after the campaigns took place. Before the campaigns were conducted, there were already differences in all effect criteria and folic acid use between women of different educational levels, mostly in favour of women with a high level of education. Although both educational campaigns appeared to have a positive impact on all outcome criteria, they failed to reduce the existing differences in these outcome criteria between women of different educational levels. Folic acid use can be promoted effectively by mass media campaigns, certainly in a large group of women with no prior knowledge of the health benefits associated with periconceptional folic acid use. However, in order to achieve more equal health outcomes among women of low and high SES, it seems that more tailored interventions for women of low SES are needed.

  9. Minimum protein intake for the preterm neonate determined by protein and amino acid kinetics.

    PubMed

    Zello, Gordon A; Menendez, Cesar E; Rafii, Mahroukh; Clarke, Ruth; Wykes, Linda J; Ball, Ronald O; Pencharz, Paul B

    2003-02-01

    Lower limits of protein needs in prematurely born neonates have not been adequately studied, yet providing protein in amounts maximizing accretion without excess is a goal in these infants' nutritional care. We hypothesized that with the use of amino acid oxidation methodology, it would be possible to define minimum protein requirement. Our objective was to investigate protein kinetics during short-term changes in protein intake by measurement of nitrogen balance and amino acid flux and oxidation using [(15)N]glycine, [(13)C]phenylalanine, and [(13)C]leucine tracers. Protein kinetics were examined in 21 preterm infants (gestational age: 29 +/- 3 wk; birth weight: 1091 +/- 324 g) at five protein intakes (1.0, 1.5, 2.0, 2.5, and 3.0 g x kg(-1) x d(-1)) with 1 d of adaptation to the test intakes. From nitrogen balance data, a protein need of 0.74 g x kg(-1 x -1) was estimated to achieve zero balance. For all three amino acids, flux and oxidation estimates were not different across protein intakes. Whole-body protein synthesis and breakdown estimates from [(15)N]ammonia data were 14.6 +/- 3.4 and 14.4 +/- 4.1 g x kg(-1) x d(-1), respectively. Glycine flux (680 +/- 168 micromol x kg(-1) x h(-1)) was greater than leucine flux (323 +/- 115 micromol x kg(-1) x h(-1)), which was greater than phenylalanine flux (84.3 +/- 35.2 micromol x kg(-1) x h(-1)). Leucine oxidation (36.7 +/- 15.6 micromol x kg(-1) x h(-1)) was also greater than phenylalanine oxidation (6.64 +/- 4.41 micromol x kg(-1) x h(-1)). Infants in our study were able to adapt to short-term changes in protein intake with little consequence to the overall whole-body protein economy, as measured by the three test amino acids.

  10. Stimulation of protein synthesis by phosphatidic acid in rat cardiomyocytes.

    PubMed

    Xu, Y J; Yau, L; Yu, L P; Elimban, V; Zahradka, P; Dhalla, N S

    1996-12-13

    Phosphatidic acid (PA) was observed to stimulate protein synthesis in adult cardiomyocytes in a time- and concentration-dependent manner. The maximal stimulation in protein synthesis (142 +/- 12% vs 100% as the control) was achieved at 10 microM PA within 60 min and was inhibited by actinomycin D (107 +/- 4% of the control) or cycloheximide (105 +/- 6% of the control). The increase in protein synthesis due to PA was attenuated or abolished by preincubation of cardiomyocytes with a tyrosine kinase inhibitor, genistein (94 +/- 9% of the control), phospholipase C inhibitors 2-nitro-4-carboxyphenyl N,N-diphenyl carbamate or carbon-odithioic acid O-(octahydro-4,7-methanol-1H-inden-5-yl (101 +/- 6 and 95 +/- 5% of the control, respectively), protein kinase C inhibitors staurosporine or polymyxin B (109 +/- 3 and 93 +/- 3% of the control), and chelators of extracellular and intracellular free Ca2+ EGTA or BAPTA/AM (103 +/- 6 and 95 +/- 6% of the control, respectively). PA at different concentrations (0.1 to 100 microM) also caused phosphorylation of a cell surface protein of approximately 24 kDa. In addition, mitogen-activated protein kinase was stimulated by PA in a concentration-dependent manner; maximal stimulation (217 +/- 6% of the control) was seen at 10 microM PA. These data suggest that PA increases protein synthesis in adult rat cardiomyocytes and thus may play an important role in the development of cardiac hypertrophy.

  11. [Photochemistry and UV Spectroscopy of Proteins and Nucleic Acids].

    PubMed

    Wierzchowski, Kazimierz Lech

    2015-01-01

    The article presents a short history of David Shugar studies in the field of photochemistry and UV spectroscopy of proteins and nucleic acids, carried out since the late 1940s. to the beginning of the 1970s. of the 20th century, with some references to the state of related research in those days.

  12. Increased isoprostane levels in oleic acid-induced lung injury

    SciTech Connect

    Ono, Koichi; Koizumi, Tomonobu; Tsushima, Kenji; Yoshikawa, Sumiko; Yokoyama, Toshiki; Nakagawa, Rikimaru; Obata, Toru

    2009-10-16

    The present study was performed to examine a role of oxidative stress in oleic acid-induced lung injury model. Fifteen anesthetized sheep were ventilated and instrumented with a lung lymph fistula and vascular catheters for blood gas analysis and measurement of isoprostanes (8-epi prostaglandin F2{alpha}). Following stable baseline measurements, oleic acid (0.08 ml/kg) was administered and observed 4 h. Isoprostane was measured by gas chromatography mass spectrometry with the isotope dilution method. Isoprostane levels in plasma and lung lymph were significantly increased 2 h after oleic acid administration and then decreased at 4 h. The percent increases in isoprostane levels in plasma and lung lymph at 2 h were significantly correlated with deteriorated oxygenation at the same time point, respectively. These findings suggest that oxidative stress is involved in the pathogenesis of the pulmonary fat embolism-induced acute lung injury model in sheep and that the increase relates with the deteriorated oxygenation.

  13. Effect of increased protein intake on renal acid load and renal hemodynamic responses.

    PubMed

    Teunissen-Beekman, Karianna F M; Dopheide, Janneke; Geleijnse, Johanna M; Bakker, Stephan J L; Brink, Elizabeth J; de Leeuw, Peter W; van Baak, Marleen A

    2016-03-01

    Increased protein intake versus maltodextrin intake for 4 weeks lowers blood pressure. Concerns exist that high-protein diets reduce renal function. Effects of acute and 4-week protein intake versus maltodextrin intake on renal acid load, glomerular filtration rate and related parameters were compared in this study. Seventy-nine overweight individuals with untreated elevated blood pressure and normal kidney function were randomized to consume a mix of protein isolates (60 g/day) or maltodextrin (60 g/day) for 4 weeks in energy balance. Twenty-four-hour urinary potential renal acid load (uPRAL) was compared between groups. A subgroup (maltodextrin N = 27, protein mix N = 25) participated in extra test days investigating fasting levels and postprandial effects of meals supplemented with a moderate protein- or maltodextrin-load on glomerular filtration rate, effective renal plasma flow, plasma renin, aldosterone, pH, and bicarbonate. uPRAL was significantly higher in the protein group after 4 weeks (P ≤ 0.001). Postprandial filtration fraction decreased further after the protein-supplemented breakfast than after the maltodextrin-supplemented breakfast after 4 weeks of supplementation (P ≤ 0.001). Fasting and postprandial levels of glomerular filtration rate, effective renal plasma flow, renin, aldosterone, angiotensin-converting enzyme, pH and bicarbonate did not differ between groups. In conclusion, 4 weeks on an increased protein diet (25% of energy intake) increased renal acid load, but did not affect renal function. Postprandial changes, except for filtration fraction, also did not differ between groups. These data suggest that a moderate increase in protein intake by consumption of a protein mix for 4 weeks causes no (undesirable) effects on kidney function in overweight and obese individuals with normal kidney function. PMID:26997623

  14. Effect of increased protein intake on renal acid load and renal hemodynamic responses.

    PubMed

    Teunissen-Beekman, Karianna F M; Dopheide, Janneke; Geleijnse, Johanna M; Bakker, Stephan J L; Brink, Elizabeth J; de Leeuw, Peter W; van Baak, Marleen A

    2016-03-01

    Increased protein intake versus maltodextrin intake for 4 weeks lowers blood pressure. Concerns exist that high-protein diets reduce renal function. Effects of acute and 4-week protein intake versus maltodextrin intake on renal acid load, glomerular filtration rate and related parameters were compared in this study. Seventy-nine overweight individuals with untreated elevated blood pressure and normal kidney function were randomized to consume a mix of protein isolates (60 g/day) or maltodextrin (60 g/day) for 4 weeks in energy balance. Twenty-four-hour urinary potential renal acid load (uPRAL) was compared between groups. A subgroup (maltodextrin N = 27, protein mix N = 25) participated in extra test days investigating fasting levels and postprandial effects of meals supplemented with a moderate protein- or maltodextrin-load on glomerular filtration rate, effective renal plasma flow, plasma renin, aldosterone, pH, and bicarbonate. uPRAL was significantly higher in the protein group after 4 weeks (P ≤ 0.001). Postprandial filtration fraction decreased further after the protein-supplemented breakfast than after the maltodextrin-supplemented breakfast after 4 weeks of supplementation (P ≤ 0.001). Fasting and postprandial levels of glomerular filtration rate, effective renal plasma flow, renin, aldosterone, angiotensin-converting enzyme, pH and bicarbonate did not differ between groups. In conclusion, 4 weeks on an increased protein diet (25% of energy intake) increased renal acid load, but did not affect renal function. Postprandial changes, except for filtration fraction, also did not differ between groups. These data suggest that a moderate increase in protein intake by consumption of a protein mix for 4 weeks causes no (undesirable) effects on kidney function in overweight and obese individuals with normal kidney function.

  15. Auxin acts independently of DELLA proteins in regulating gibberellin levels.

    PubMed

    Reid, James B; Davidson, Sandra E; Ross, John J

    2011-03-01

    Shoot elongation is a vital process for plant development and productivity, in both ecological and economic contexts. Auxin and bioactive gibberellins (GAs), such as GA1, play critical roles in the control of elongation, along with environmental and endogenous factors, including other hormones such as the brassinosteroids. The effect of auxins, such as indole-3-acetic acid (IAA), is at least in part mediated by its effect on GA metabolism, since auxin up-regulates biosynthesis genes such as GA 3-oxidase and GA 20-oxidase and down regulates GA catabolism genes such as GA 2-oxidases, leading to elevated levels of bioactive GA 1. In our recent paper, we have provided evidence that this action of IAA is largely independent of DELLA proteins, the negative regulators of GA action, since the auxin effects are still present in the DELLA-deficient la cry-s genotype of pea. This was a crucial issue to resolve, since like auxin, the DELLAs also promote GA 1 synthesis and inhibit its deactivation. DELLAs are deactivated by GA, and thereby mediate a feedback system by which bioactive GA regulates its own level. However, our recent results, in themselves, do not show the generality of the auxin-GA relationship across species and phylogenetic groups or across different tissue types and responses. Further, they do not touch on the ecological benefits of the auxin-GA interaction. These issues are discussed below as well as the need for the development of suitable experimental systems to allow this process to be examined. PMID:21358281

  16. Relationship between Uric Acid Level and Achievement Motivation. Final Report.

    ERIC Educational Resources Information Center

    Mueller, Ernst F.; French, John R. P., Jr.

    In an investigation of the relationship of uric acid (a metabolic end product) to achievement, this study hypothesized that a person's serum urate level (a factor often associated with gout) is positively related to achievement need as well as indicators of actual achievement. (Speed of promotion and number of yearly publications were chosen as…

  17. Dietary protein level and performance of growing Baladi kids.

    PubMed

    Abdelrahman, M M; Aljumaah, R S

    2014-01-01

    A study was conducted to evaluate the effect of feeding different levels of protein to black Baladi breed kids. Weanling Baladi kids (n=18; 75 to 90 days old) were selected and individually housed at our experimental farm. Kids were divided randomly to one of the three treatments for 12 weeks. The three dietary treatments were: T1: control ration, formulated according to NRC to cover the protein (level 1) and other nutrients requirements. T2: ration formulated to cover only 75% of protein (level 2) recommended by NRC. T3: control diet + 2.4 g undegradable methionine (Smartamine®)/day/kid (level 3). Feed intake, initial and monthly body weights were recorded. Blood samples were collected monthly and analyzed for metabolites and Co, Zn and Cu levels. Decreasing the dietary level of protein (T2) negatively affected (P<0.05) the total live weight gain, average daily gain and feed conversion ratio when compared with the control and T3 groups. Moreover, treatment, time and time × treatment caused a significant change on Co concentration in blood serum with higher value at the end of the experiment. Treatments had a significant effect (P<0.05) on blood serum cholesterol and protein levels. Undegradable methionine supplementation (T3) significantly increased longissimus dorsi weight, fat thickness and omental fat%. In conclusion, feeding Baladi kids below the NRC requirements of protein negatively affect the growth performance and feed efficiency. The recommended protein level by NRC for growing kids cover the requirements of growing black Baladi kids for maximum growth and productivity.

  18. Dietary protein level and performance of growing Baladi kids

    PubMed Central

    Abdelrahman, M. M.; Aljumaah, R. S.

    2014-01-01

    A study was conducted to evaluate the effect of feeding different levels of protein to black Baladi breed kids. Weanling Baladi kids (n=18; 75 to 90 days old) were selected and individually housed at our experimental farm. Kids were divided randomly to one of the three treatments for 12 weeks. The three dietary treatments were: T1: control ration, formulated according to NRC to cover the protein (level 1) and other nutrients requirements. T2: ration formulated to cover only 75% of protein (level 2) recommended by NRC. T3: control diet + 2.4 g undegradable methionine (Smartamine®)/day/kid (level 3). Feed intake, initial and monthly body weights were recorded. Blood samples were collected monthly and analyzed for metabolites and Co, Zn and Cu levels. Decreasing the dietary level of protein (T2) negatively affected (P<0.05) the total live weight gain, average daily gain and feed conversion ratio when compared with the control and T3 groups. Moreover, treatment, time and time × treatment caused a significant change on Co concentration in blood serum with higher value at the end of the experiment. Treatments had a significant effect (P<0.05) on blood serum cholesterol and protein levels. Undegradable methionine supplementation (T3) significantly increased longissimus dorsi weight, fat thickness and omental fat%. In conclusion, feeding Baladi kids below the NRC requirements of protein negatively affect the growth performance and feed efficiency. The recommended protein level by NRC for growing kids cover the requirements of growing black Baladi kids for maximum growth and productivity. PMID:27175130

  19. Leukocyte protease binding to nucleic acids promotes nuclear localization and cleavage of nucleic acid binding proteins.

    PubMed

    Thomas, Marshall P; Whangbo, Jennifer; McCrossan, Geoffrey; Deutsch, Aaron J; Martinod, Kimberly; Walch, Michael; Lieberman, Judy

    2014-06-01

    Killer lymphocyte granzyme (Gzm) serine proteases induce apoptosis of pathogen-infected cells and tumor cells. Many known Gzm substrates are nucleic acid binding proteins, and the Gzms accumulate in the target cell nucleus by an unknown mechanism. In this study, we show that human Gzms bind to DNA and RNA with nanomolar affinity. Gzms cleave their substrates most efficiently when both are bound to nucleic acids. RNase treatment of cell lysates reduces Gzm cleavage of RNA binding protein targets, whereas adding RNA to recombinant RNA binding protein substrates increases in vitro cleavage. Binding to nucleic acids also influences Gzm trafficking within target cells. Preincubation with competitor DNA and DNase treatment both reduce Gzm nuclear localization. The Gzms are closely related to neutrophil proteases, including neutrophil elastase (NE) and cathepsin G. During neutrophil activation, NE translocates to the nucleus to initiate DNA extrusion into neutrophil extracellular traps, which bind NE and cathepsin G. These myeloid cell proteases, but not digestive serine proteases, also bind DNA strongly and localize to nuclei and neutrophil extracellular traps in a DNA-dependent manner. Thus, high-affinity nucleic acid binding is a conserved and functionally important property specific to leukocyte serine proteases. Furthermore, nucleic acid binding provides an elegant and simple mechanism to confer specificity of these proteases for cleavage of nucleic acid binding protein substrates that play essential roles in cellular gene expression and cell proliferation.

  20. Dietary docosahexaenoic acid and docosapentaenoic acid ameliorate amyloid-beta and tau pathology via a mechanism involving presenilin 1 levels.

    PubMed

    Green, Kim N; Martinez-Coria, Hilda; Khashwji, Hasan; Hall, Eileen B; Yurko-Mauro, Karin A; Ellis, Lorie; LaFerla, Frank M

    2007-04-18

    The underlying cause of sporadic Alzheimer disease (AD) is unknown, but a number of environmental and genetic factors are likely to be involved. One environmental factor that is increasingly being recognized as contributing to brain aging is diet, which has evolved markedly over modern history. Here we show that dietary supplementation with docosahexaenoic acid (DHA), an n-3 polyunsaturated fatty acid, in the 3xTg-AD mouse model of AD reduced the intraneuronal accumulation of both amyloid-beta (Abeta) and tau. In contrast, combining DHA with n-6 fatty acids, either arachidonic acid or docosapentaenoic acid (DPAn-6), diminished the efficacy of DHA over a 12 month period. Here we report the novel finding that the mechanism accounting for the reduction in soluble Abeta was attributable to a decrease in steady-state levels of presenilin 1, and not to altered processing of the amyloid precursor protein by either the alpha- or beta-secretase. Furthermore, the presence of DPAn-6 in the diet reduced levels of early-stage phospho-tau epitopes, which correlated with a reduction in phosphorylated c-Jun N-terminal kinase, a putative tau kinase. Collectively, these results suggest that DHA and DPAn-6 supplementations could be a beneficial natural therapy for AD.

  1. Meal composition and plasma amino acid ratios: Effect of various proteins or carbohydrates, and of various protein concentrations

    NASA Technical Reports Server (NTRS)

    Yokogoshi, Hidehiko; Wurtman, Richard J.

    1986-01-01

    The effects of meals containing various proteins and carbohydrates, and of those containing various proportions of protein (0 percent to 20 percent of a meal, by weight) or of carbohydrate (0 percent to 75 percent), on plasma levels of certain large neutral amino acids (LNAA) in rats previously fasted for 19 hours were examined. Also the plasma tryptophan ratios (the ratio of the plasma trytophan concentration to the summed concentrations of the other large neutral amino acids) and other plasma amino acid ratios were calculated. (The plasma tryptophan ratio has been shown to determine brain tryptophan levels and, thereby, to affect the synthesis and release of the neurotransmitter serotonin). A meal containing 70 percent to 75 percent of an insulin-secreting carbohydrate (dextrose or dextrin) increased plasma insulin levels and the tryptophan ratio; those containing 0 percent or 25 percent carbohydrate failed to do so. Addition of as little as 5 percent casein to a 70 percent carbohydrate meal fully blocked the increase in the plasma tryptophan ratio without affecting the secretion of insulin - probably by contributing much larger quantities of the other LNAA than of tryptophan to the blood. Dietary proteins differed in their ability to suppress the carbohydrate-induced rise in the plasma tryptophan ratio. Addition of 10 percent casein, peanut meal, or gelatin fully blocked this increase, but lactalbumin failed to do so, and egg white did so only partially. (Consumption of the 10 percent gelatin meal also produced a major reduction in the plasma tyrosine ratio, and may thereby have affected brain tyrosine levels and catecholamine synthesis.) These observations suggest that serotonin-releasing neurons in brains of fasted rats are capable of distinguishing (by their metabolic effects) between meals poor in protein but rich in carbohydrates that elicit insulin secretion, and all other meals. The changes in brain serotonin caused by carbohydrate-rich, protein

  2. A conserved patch of hydrophobic amino acids modulates Myb activity by mediating protein-protein interactions.

    PubMed

    Dukare, Sandeep; Klempnauer, Karl-Heinz

    2016-07-01

    The transcription factor c-Myb plays a key role in the control of proliferation and differentiation in hematopoietic progenitor cells and has been implicated in the development of leukemia and certain non-hematopoietic tumors. c-Myb activity is highly dependent on the interaction with the coactivator p300 which is mediated by the transactivation domain of c-Myb and the KIX domain of p300. We have previously observed that conservative valine-to-isoleucine amino acid substitutions in a conserved stretch of hydrophobic amino acids have a profound effect on Myb activity. Here, we have explored the function of the hydrophobic region as a mediator of protein-protein interactions. We show that the hydrophobic region facilitates Myb self-interaction and binding of the histone acetyl transferase Tip60, a previously identified Myb interacting protein. We show that these interactions are affected by the valine-to-isoleucine amino acid substitutions and suppress Myb activity by interfering with the interaction of Myb and the KIX domain of p300. Taken together, our work identifies the hydrophobic region in the Myb transactivation domain as a binding site for homo- and heteromeric protein interactions and leads to a picture of the c-Myb transactivation domain as a composite protein binding region that facilitates interdependent protein-protein interactions of Myb with regulatory proteins.

  3. Probing the structural dynamics of proteins and nucleic acids with optical tweezers.

    PubMed

    Ritchie, Dustin B; Woodside, Michael T

    2015-10-01

    Conformational changes are an essential feature of most molecular processes in biology. Optical tweezers have emerged as a powerful tool for probing conformational dynamics at the single-molecule level because of their high resolution and sensitivity, opening new windows on phenomena ranging from folding and ligand binding to enzyme function, molecular machines, and protein aggregation. By measuring conformational changes induced in a molecule by forces applied by optical tweezers, new insight has been gained into the relationship between dynamics and function. We discuss recent advances from studies of how structure forms in proteins and RNA, including non-native structures, fluctuations in disordered proteins, and interactions with chaperones assisting native folding. We also review the development of assays probing the dynamics of complex protein-nucleic acid and protein-protein assemblies that reveal the dynamic interactions between biomolecular machines and their substrates.

  4. Puget Sound acidity levels drop after ASARCO shutdown

    SciTech Connect

    Not Available

    1987-07-01

    The levels of acidity in Puget Sound region rainfall have decreased significantly since the shutdown of the ASARCO copper smelter in Tacoma, Washington, according to a study funded by the US Environmental Protection Agency. Results indicate that sulfate and hydrogen ion concentrations obtained from samples taken before the closure were significantly different than those collected after the shutdown. Rainwater samples collected downwind during smelter operation were also significantly different from those collected upwind. Sulfur dioxide is considered to be one of the principal contributors to acid rain. The smelter was a major source of sulfur dioxide emissions in the Puget Sound region before it shut down in March 1985.

  5. Analysis of soybean root proteins affected by gibberellic acid treatment under flooding stress.

    PubMed

    Oh, Myeong Won; Nanjo, Yohei; Komatsu, Setsuko

    2014-01-01

    Flooding is a serious abiotic stress for soybean because it restricts growth and reduces grain yields. To investigate the effect of gibberellic acid (GA) on soybean under flooding stress, root proteins were analyzed using a gel-free proteomic technique. Proteins were extracted from the roots of 4-days-old soybean seedlings exposed to flooding stress in the presence and absence of exogenous GA3 for 2 days. A total of 307, 324, and 250 proteins were identified from untreated, and flooding-treated soybean seedlings without or with GA3, respectively. Secondary metabolism- and cell-related proteins, and proteins involved in protein degradation/synthesis were decreased by flooding stress; however, the levels of these proteins were restored by GA3 supplementation under flooding. Fermentation- and cell wall-related proteins were not affected by GA3 supplementation. Furthermore, putative GA-responsive proteins, which were identified by the presence of a GA-responsive element in the promoter region, were less abundant by flooding stress; however, these proteins were more abundant by GA3 supplementation under flooding. Taken together, these results suggest that GA3 affects the abundance of proteins involved in secondary metabolism, cell cycle, and protein degradation/synthesis in soybeans under flooding stress. PMID:24702262

  6. Plasma protein carbonyl levels and breast cancer risk.

    PubMed

    Rossner, Pavel; Terry, Mary Beth; Gammon, Marilie D; Agrawal, Meenakshi; Zhang, Fang Fang; Ferris, Jennifer S; Teitelbaum, Susan L; Eng, Sybil M; Gaudet, Mia M; Neugut, Alfred I; Santella, Regina M

    2007-01-01

    To study the role of oxidative stress in breast cancer risk, we analysed plasma levels of protein carbonyls in 1050 cases and 1107 controls. We found a statistically significant trend in breast cancer risk in relation to increasing quartiles of plasma protein carbonyl levels (OR = 1.2, 95% CI = 0.9-1.5; OR = 1.5, 95% CI = 1.2-2.0; OR = 1.6, 95% CI = 1.2-2.1, for the 2(nd), 3(rd) and 4(th) quartile relative to the lowest quartile, respectively, P for trend = 0.0001). The increase in risk was similar for younger (<50 years) and older women, more pronounced among women with higher physical activity levels (0.7 hrs/week for 4(th) quartile versus lowest quartile OR = 2.0, 95% CI = 1.4-3.0), higher alcohol consumption (> or = 15 grams/day for 4(th) quartile versus lowest quartile OR = 2.3, 95% CI = 1.1-4.7), and hormone replacement therapy use (HRT, OR = 2.6, 95% CI = 1.6-4.4 for 4(th) quartile versus lowest quartile). The multiplicative interaction terms were statistically significant only for physical activity and HRT. The positive association between plasma protein carbonyl levels and breast cancer risk was also observed when the analysis was restricted to women who had not received chemotherapy or radiation therapy prior to blood collection. Among controls, oxidized protein levels significantly increased with cigarette smoking and higher fruit and vegetable consumption, and decreased with alcohol consumption >30 grams per day. Women with higher levels of plasma protein carbonyl and urinary 15F(2t)-isoprostane had an 80% increase in breast cancer risk (OR = 1.8, 95% CI = 1.2-2.6) compared to women with levels below the median for both markers of oxidative stress. In summary, our results suggest that increased plasma protein carbonyl levels may be associated with breast cancer risk.

  7. The Mediterranean diet: Effects on proteins that mediate fatty acid metabolism in the colon

    PubMed Central

    Djuric, Zora

    2012-01-01

    A Mediterranean diet appears to have health benefits in many domains of human health, mediated perhaps by its anti-inflammatory effects. Metabolism of fatty acids and subsequent eicosanoid production is a key mechanism by which a Mediterranean diet can exert anti-inflammatory effects. Both dietary fatty acids and fatty acid metabolism determine fatty acid availability for cyclooxygenase- and lipoxygenase-dependent production of eicosanoids, namely prostaglandins and leukotrienes. In dietary intervention studies and in observational studies of the Mediterranean diet, blood levels of fatty acids do reflect dietary intakes but are attenuated. Small differences in fatty acid levels, however, appear to be important, especially when exposures occur over long periods of time. This review summarizes how fat intakes from a Greek-style Mediterranean diet can be expected to affect fatty acid metabolizing proteins, with an emphasis on the metabolic pathways that lead to the formation of proinflammatory eicosanoids. The proteins involved in these pathways are ripe for investigation using proteomic approaches and may be targets for colon cancer prevention. PMID:22133197

  8. In Silico Classification of Proteins from Acidic and Neutral Cytoplasms

    PubMed Central

    Fang, Yaping; Middaugh, C. Russell; Fang, Jianwen

    2012-01-01

    Protein acidostability is a common problem in biopharmaceutical and other industries. However, it remains a great challenge to engineer proteins for enhanced acidostability because our knowledge of protein acidostabilization is still very limited. In this paper, we present a comparative study of proteins from bacteria with acidic (AP) and neutral cytoplasms (NP) using an integrated statistical and machine learning approach. We construct a set of 393 non-redundant AP-NP ortholog pairs and calculate a total of 889 sequence based features for these proteins. The pairwise alignments of these ortholog pairs are used to build a residue substitution propensity matrix between APs and NPs. We use Gini importance provided by the Random Forest algorithm to rank the relative importance of these features. A scoring function using the 10 most significant features is developed and optimized using a hill climbing algorithm. The accuracy of the score function is 86.01% in predicting AP-NP ortholog pairs and is 76.65% in predicting non-ortholog AP-NP pairs, suggesting that there are significant differences between APs and NPs which can be used to predict relative acidostability of proteins. The overall trends uncovered in the study can be used as general guidelines for designing acidostable proteins. To best of our knowledge, this work represents the first systematic comparative study of the acidostable proteins and their non-acidostable orthologs. PMID:23049817

  9. Soy protein isolate molecular level contributions to bulk adhesive properties

    NASA Astrophysics Data System (ADS)

    Shera, Jeanne Norton

    Increasing environmental awareness and the recognized health hazards of formaldehyde-based resins has prompted a strong demand for environmentally-responsible adhesives for wood composites. Soy protein-based adhesives have been shown to be commercially viable with 90-day shelf stability and composite physical properties comparable to those of commercial formaldehyde-based particleboards. The main research focus is to isolate and characterize the molecular level features in soy protein isolate responsible for providing mechanical properties, storage stability, and water resistance during adhesive formulation, processing, and wood composite fabrication. Commercial composite board will be reviewed to enhance our understanding of the individual components and processes required for particleboard production. The levels of protein structure will be defined and an overview of current bio-based technology will be presented. In the process, the logic for utilizing soy protein as a sole binder in the adhesive will be reinforced. Variables such as adhesive components, pH, divalent ions, blend aging, protein molecular weight, formulation solids content, and soy protein functionalization will relate the bulk properties of soy protein adhesives to the molecular configuration of the soybean protein. This work has demonstrated that when intermolecular beta-sheet interactions and protein long-range order is disrupted, viscosity and mechanical properties decrease. Storage stability can be maintained through the stabilization of intermolecular beta-sheet interactions. When molecular weight is reduced through enzymatic digestion, long-range order is disrupted and viscosity and mechanical properties decrease accordingly. Processibility and physical properties must be balanced to increase solids while maintaining low viscosity, desirable mechanical properties, and adequate storage stability. The structure of the soybean protein must be related to the particleboard bulk mechanical

  10. Targeted modification of storage protein content resulting in improved amino acid composition of barley grain.

    PubMed

    Sikdar, Md S I; Bowra, S; Schmidt, D; Dionisio, G; Holm, P B; Vincze, E

    2016-02-01

    C-hordein in barley and ω-gliadins in wheat are members of the prolamins protein families. Prolamins are the major component of cereal storage proteins and composed of non-essential amino acids (AA) such as proline and glutamine therefore have low nutritional value. Using double stranded RNAi silencing technology directed towards C-hordein we obtained transgenic barley lines with up to 94.7% reduction in the levels of C-hordein protein relative to the parental line. The composition of the prolamin fraction of the barley parental line cv. Golden Promise was resolved using SDS-PAGE electrophoresis, the protein band were excised and the proteins identified by quadrupole-time-of-flight mass spectrometry. Subsequent SDS-PAGE separation and analysis of the prolamin fraction of the transgenic lines revealed a reduction in the amounts of C-hordeins and increases in the content of other hordein family members. Analysis of the AA composition of the transgenic lines showed that the level of essential amino acids increased with a concomitant reduction in proline and glutamine. Both the barley C-hordein and wheat ω-gliadin genes proved successful for RNAi-gene mediated suppression of barley C-hordein level. All transgenic lines that exhibited a reduction for C-hordein showed off-target effects: the lines exhibited increased level of B/γ-hordein while D-hordein level was reduced. Furthermore, the multicopy insertions correlated negatively with silencing.

  11. How Integrated Management Strategies Promote Protein Quality of Cotton Embryos: High Levels of Soil Available N, N Assimilation and Protein Accumulation Rate

    PubMed Central

    Yang, HongKun; Meng, YaLi; Chen, BingLin; Zhang, XingYue; Wang, YouHua; Zhao, WenQing; Zhou, ZhiGuo

    2016-01-01

    Cottonseed is widely used as a source of ruminant feed and for industrial purposes. Therefore, there is a tremendous need to improve the nutritional value of cotton embryos. In this study, a conventional management (CM) and two integrated cotton management strategies (IMS1, IMS2) were performed at two soil fertility levels to study the relationships among soil N, N assimilation, embryonic protein accumulation and protein quality. The levels of proteins, essential amino acids, and semi-essential amino acids, especially those of glutamate, lysine, and methionine, were higher in IMS1 and IMS2 embryos than in CM embryos. These changes were significantly positively correlated with the soil-available N content, glutamine synthetase activity and peak value of protein accumulation rate and were negatively correlated with the free amino acid level. These results illustrated that integrated management strategies, especially the rates and timing of N application, raise the level of soil available N, which is beneficial for N assimilation in developing cotton embryos. The protein content was limited by the rate of protein accumulation rather than by the free amino acid content. The combination of target yield fertilization, a growth-driven N application schedule, a high plant density and the seedling raising with bio-organic fertilizer can substantially improve protein quality in cotton embryos, especially at a soil with low soil organic matter and total nitrogen. PMID:27532007

  12. How Integrated Management Strategies Promote Protein Quality of Cotton Embryos: High Levels of Soil Available N, N Assimilation and Protein Accumulation Rate.

    PubMed

    Yang, HongKun; Meng, YaLi; Chen, BingLin; Zhang, XingYue; Wang, YouHua; Zhao, WenQing; Zhou, ZhiGuo

    2016-01-01

    Cottonseed is widely used as a source of ruminant feed and for industrial purposes. Therefore, there is a tremendous need to improve the nutritional value of cotton embryos. In this study, a conventional management (CM) and two integrated cotton management strategies (IMS1, IMS2) were performed at two soil fertility levels to study the relationships among soil N, N assimilation, embryonic protein accumulation and protein quality. The levels of proteins, essential amino acids, and semi-essential amino acids, especially those of glutamate, lysine, and methionine, were higher in IMS1 and IMS2 embryos than in CM embryos. These changes were significantly positively correlated with the soil-available N content, glutamine synthetase activity and peak value of protein accumulation rate and were negatively correlated with the free amino acid level. These results illustrated that integrated management strategies, especially the rates and timing of N application, raise the level of soil available N, which is beneficial for N assimilation in developing cotton embryos. The protein content was limited by the rate of protein accumulation rather than by the free amino acid content. The combination of target yield fertilization, a growth-driven N application schedule, a high plant density and the seedling raising with bio-organic fertilizer can substantially improve protein quality in cotton embryos, especially at a soil with low soil organic matter and total nitrogen. PMID:27532007

  13. How Integrated Management Strategies Promote Protein Quality of Cotton Embryos: High Levels of Soil Available N, N Assimilation and Protein Accumulation Rate.

    PubMed

    Yang, HongKun; Meng, YaLi; Chen, BingLin; Zhang, XingYue; Wang, YouHua; Zhao, WenQing; Zhou, ZhiGuo

    2016-01-01

    Cottonseed is widely used as a source of ruminant feed and for industrial purposes. Therefore, there is a tremendous need to improve the nutritional value of cotton embryos. In this study, a conventional management (CM) and two integrated cotton management strategies (IMS1, IMS2) were performed at two soil fertility levels to study the relationships among soil N, N assimilation, embryonic protein accumulation and protein quality. The levels of proteins, essential amino acids, and semi-essential amino acids, especially those of glutamate, lysine, and methionine, were higher in IMS1 and IMS2 embryos than in CM embryos. These changes were significantly positively correlated with the soil-available N content, glutamine synthetase activity and peak value of protein accumulation rate and were negatively correlated with the free amino acid level. These results illustrated that integrated management strategies, especially the rates and timing of N application, raise the level of soil available N, which is beneficial for N assimilation in developing cotton embryos. The protein content was limited by the rate of protein accumulation rather than by the free amino acid content. The combination of target yield fertilization, a growth-driven N application schedule, a high plant density and the seedling raising with bio-organic fertilizer can substantially improve protein quality in cotton embryos, especially at a soil with low soil organic matter and total nitrogen.

  14. Protein metabolism in growing pigs fed corn or cassava peel based diets containing graded protein levels.

    PubMed

    Tewe, O O

    1985-05-01

    Sixty-four Large White cross Landrace weanling pigs were randomly allotted to eight treatments in a two by four factorial arrangement. The two dietary variables were cassava peel (0 and 40 per cent) and crude protein (20, 15, 10 and 5 per cent). Total serum protein concentration was significantly (P less than 0.01) reduced by protein deficiency and by its interaction with cassava peel. The multiple coefficient of determination (R2) showed that protein intake was the primary factor determining changes in serum protein. R2 values for cyanide intake (independent variable) on serum protein (dependent variable) increased from day 30 to 90 of the trial. Serum urea was increased on the 5 per cent protein diets on days 60 and 90 of the trial. The R2 values for cyanide and protein intake on serum urea concentration increased from day 30 to day 90 of the trial. Serum creatinine increased (P less than 0.05) on the 5 per cent protein diet on day 90 of the trial. The R2 value for the effects of protein intake on serum creatinine was higher than for cyanide intake on days 30 and 90. The results confirm the progressive and pronounced effects of long term cyanide intake on serum nitrogenous metabolites in pigs consuming between 110 and 120 ppm hydrocyanic acid, especially in diets containing 10 per cent or less protein. PMID:2989987

  15. Single-Cell Protein Production by the Acid-Tolerant Fungus Scytalidium acidophilum from Acid Hydrolysates of Waste Paper †

    PubMed Central

    Ivarson, K. C.; Morita, H.

    1982-01-01

    The bioconversion of waste paper to single-cell protein at pH <1 by Scytalidium acidophilum is described. Waste paper pretreated with 72% H2SO4 at 4°C was diluted with water to a pH of <0.1 and hydrolyzed. This yielded an adequate sugar-containing substrate for the growth of the fungus. A total of 97% of the sugars (glucose, galactose, mannose, xylose, arabinose) in the hydrolysates were converted to cell biomass. Microbial contamination was not observed. Based on the sugars consumed, S. acidophilum produced higher yields in shake cultures than many other Fungi Imperfecti. In aerated cultures, productivity increased, and yields of 43 to 46% containing 44 to 47% crude protein were obtained. This compares favorably with Candida utilis, a yeast used commercially to produce single-cell protein. The chemical constituents and the essential amino acids of the fungal cells were similar to those of other fungi. The nucleic acid content was characteristic of microbes containing low levels of nucleic acid. The advantages of using S. acidophilum for single-cell protein production are discussed. PMID:16345970

  16. High Level Waste System Impacts from Acid Dissolution of Sludge

    SciTech Connect

    KETUSKY, EDWARD

    2006-04-20

    This research evaluates the ability of OLI{copyright} equilibrium based software to forecast Savannah River Site High Level Waste system impacts from oxalic acid dissolution of Tank 1-15 sludge heels. Without further laboratory and field testing, only the use of oxalic acid can be considered plausible to support sludge heel dissolution on multiple tanks. Using OLI{copyright} and available test results, a dissolution model is constructed and validated. Material and energy balances, coupled with the model, identify potential safety concerns. Overpressurization and overheating are shown to be unlikely. Corrosion induced hydrogen could, however, overwhelm the tank ventilation. While pH adjustment can restore the minimal hydrogen generation, resultant precipitates will notably increase the sludge volume. OLI{copyright} is used to develop a flowsheet such that additional sludge vitrification canisters and other negative system impacts are minimized. Sensitivity analyses are used to assess the processability impacts from variations in the sludge/quantities of acids.

  17. Fatty acid-binding site environments of serum vitamin D-binding protein and albumin are different

    PubMed Central

    Swamy, Narasimha; Ray, Rahul

    2008-01-01

    Vitamin D-binding protein (DBP) and albumin (ALB) are abundant serum proteins and both possess high-affinity binding for saturated and unsaturated fatty acids. However, certain differences exist. We surmised that in cases where serum albumin level is low, DBP presumably can act as a transporter of fatty acids. To explore this possibility we synthesized several alkylating derivatives of 14C-palmitic acid to probe the fatty acid binding pockets of DBP and ALB. We observed that N-ethyl-5-phenylisooxazolium-3′-sulfonate-ester (WRK ester) of 14C-palmitic acid specifically labeled DBP; but p-nitrophenyl- and N-hydroxysuccinimidyl-esters failed to do so. However, p-nitrophenyl ester of 14C-palmitic acid specifically labeled bovine ALB, indicating that the micro-environment of the fatty acid-binding domains of DBP and ALB may be different; and DBP may not replace ALB as a transporter of fatty acids. PMID:18374965

  18. Effect of Maillard browning reaction on protein utilization and plasma amino acid response by rainbow trout (Salmo gairdneri).

    PubMed

    Plakas, S M; Lee, T C; Wolke, R E; Meade, T L

    1985-12-01

    The effect of the Maillard browning reaction in the diet of rainbow trout (Salmo gairdneri) on growth and amino acid availability was investigated. Chemical and enzymatic hydrolysis methods were applied for the detection of the losses of amino acids in a model protein browning system. Arginine and lysine exhibited the greatest losses in the mixture of fish protein isolate and glucose stored for 40 d at 37 degrees C. The apparent digestibility and absorption of individual amino acids, particularly lysine, was lower in trout fed browned protein than in those fed the control protein. Plasma lysine levels were significantly depressed, while the plasma levels of glucose and most other amino acids were elevated in relation to the loss in nutritive value of dietary protein after browning. The early Maillard reaction derivative of lysine, epsilon-deoxy-fructosyl-lysine, was recovered from browned protein (by using the in vitro enzymatic hydrolysis procedure) and from the plasma of trout fed browned protein. Analysis of plasma free amino acids provided an indication of lysine bioavailability and identified lysine as the first-limiting amino acid in the diets containing browned protein. PMID:3934350

  19. Altered Serum Uric Acid Level in Lichen Planus Patients

    PubMed Central

    Chakraborti, Goutam; Biswas, Rabindranath; Chakraborti, Sandip; Sen, Pradyot Kumar

    2014-01-01

    Background: Lichen planus (LP) is a common disorder whose etiopathogenesis is not clear. Recently, it has been suggested that increased reactive oxygen species (ROS) play important roles in the underlying mechanism of LP. Objectives: The principal aim of this study was to evaluate serum uric acid (UA) levels as a measure of the antioxidant defense status in LP patients. Methods: Serum UA levels were determined in 58 LP patients and 61 controls. Results: Serum UA levels were significantly decreased in patients with respect to controls. Moreover, serum UA level was decreased according to increasing duration of disease. Conclusions: The results of our study suggest that LP is associated with decrease of UA levels in serum. UA may be a potential, useful biomarker of antioxidant status in LP for elaboration of treatment strategy and monitoring. PMID:25484383

  20. Immunocytochemical staining for glial fibrillary acidic protein and the metabolism of cytoskeletal proteins in experimental allergic encephalomyelitis.

    PubMed

    Smith, M E; Somera, F P; Eng, L F

    1983-04-01

    Spinal cord sections from Lewis rats with acute experimental allergic encephalomyelitis (EAE) showed greatly increased staining of astrocytes when stained immunocytochemically for glial fibrillary acidic protein (GFAP). Fibrous processes in white matter were heavily stained early in the course of the disease when paralysis was first evident (10-12 days after injection of guinea pig spinal cord myelin), then protoplasmic astrocytes were stained in the gray matter and became more heavily stained at 20 days post-injection. The stained astrocytes were evenly distributed throughout the tissue, and did not correspond to the sites of the lesions. Spinal cord slices of control and EAE rats were incubated with [3H]amino acids, then cytoskeletal proteins were prepared in an enriched fraction, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and the protein bands counted for radioactivity. In the EAE rat all cytoskeletal proteins, including the neurofilaments, vimentin, microtubules, GFAP and actin, showed increased uptake of radioactive amino acids. Immunoprecipitation of GFAP with specific antiserum showed increased radioactivity in the complex beginning at day 10 when cellular infiltration was beginning in the EAE animals. As the disease became acute, the radioactivity in the immunoprecipitated GFAP increased, in some cases to very high levels, then by day 18 when recovery was underway, the radioactivity had fallen to normal levels. Possible agents causing metabolic activation of protein synthesis in EAE animals include stimulating substances elaborated by infiltrating lymphoid cells, and the generalized edema accompanying the demyelinative condition. The activation of GFAP protein staining and metabolism in EAE might serve as a model for the activated growth of astrocyte processes which cause the severe gliosis seen in multiple sclerosis.

  1. Protein and amino acid requirements in human nutrition.

    PubMed

    2007-01-01

    The World Health Organization and the Food and Agriculture Organization have worked to quantify the energy and nutrient needs of populations since 1949. This is the latest in a series of reports that aim to provide: updates on protein and amino acid requirements in health and disease for all age groups and pregnant and lactating mothers; recommendations on protein requirements in health and disease, including their implications for developing countries; recommendations on protein quality and labelling for worldwide use. This report provides the tools to address practical questions on the adequacy of food supplies, targets for food and nutrition policy, and labelling of protein quality. It contains specific recommendations for infant, child and adult nutrition. This report is an essential reference for those who need to determine the adequacy of population food intakes; set national food and nutrition guidelines and regulations on the protein and amino acid content of industrially processed foods; determine nutrient needs, and evaluate and ensure the adequacy of rations for vulnerable groups. The tools in this report can also be used to map and monitor global food shortages and under-nutrition through early warning systems.

  2. Tolerance to acetic acid is improved by mutations of the TATA-binding protein gene.

    PubMed

    An, Jieun; Kwon, Hyeji; Kim, Eunjung; Lee, Young Mi; Ko, Hyeok Jin; Park, Hongjae; Choi, In-Geol; Kim, Sooah; Kim, Kyoung Heon; Kim, Wankee; Choi, Wonja

    2015-03-01

    Screening a library of overexpressing mutant alleles of the TATA-binding gene SPT15 yielded two Saccharomyces cerevisiae strains (MRRC 3252 and 3253) with enhanced tolerance to acetic acid. They were also tolerant to propionic acid and hydrogen peroxide. Transcriptome profile analysis identified 58 upregulated genes and 106 downregulated genes in MRRC 3252. Stress- and protein synthesis-related transcription factors were predominantly enriched in the upregulated and downregulated genes respectively. Eight deletion mutants for some of the highly downregulated genes were acetic acid-tolerant. The level of intracellular reactive oxygen species was considerably lessened in MRRC 3252 and 3253 upon exposure to acetic acid. Metabolome profile analysis revealed that intracellular concentrations of 5 and 102 metabolites were increased and decreased, respectively, in MRRC 3252, featuring a large increase of urea and a significant decrease of amino acids. The dur1/2Δmutant, in which the urea degradation gene DUR1/2 is deleted, displayed enhanced tolerance to acetic acid. Enhanced tolerance to acetic acid was also observed on the medium containing a low concentration of amino acids. Taken together, this study identified two SPT15 alleles, nine gene deletions and low concentration of amino acids in the medium that confer enhanced tolerance to acetic acid.

  3. Fatty acid binding protein in the intestine of the chicken.

    PubMed

    Katongole, J B; March, B E

    1979-03-01

    The mucosa of the mesenteric intestine of the chicken has been found to contain a fatty acid binding protein (FABP) with a molecular weight of less than 12,400. The protein is present in the newly hatched chick before ingestion of feed and in the adult bird. When a low-fat diet is fed, the concentration of the FABP is highest in the proximal portion of the intestine and decreases posteriorly. When a high-fat diet is fed, an increase occurs in the amount of FABP in the lower section of the intestine.

  4. [Effects of exogenous nitric oxide, salicylic acid and hydrogen peroxide on free amino acid and soluble protein contents in tobacco leaves].

    PubMed

    Wei, Xiao-Hong; Wang, Li-Min; Long, Rui-Jun; Wang, Gen-Xuan

    2006-04-01

    The work focused on the effects of signal molecules of nitric oxide (NO), salicylic acid (SA) and hydrogen peroxide (H(2)O(2)) on compounds of nitric metabolites of free amino acid, soluble protein and proline in tobacco. The results indicated that NO, SA and H(2)O(2) were able to regulate the proline, free amino acid and soluble protein content in tobacco. Lower concentration of NO and H(2)O(2) raised the level of proline; while higher concentration of NO and H(2)O(2) lowered the proline, free amino acid content in tobacco. The above three signal molecules also showed similar effects on proline, free amino acid and soluble protein content in tobacco.

  5. Fatty acid binding protein 7 and n-3 poly unsaturated fatty acid supply in early rat brain development.

    PubMed

    Maximin, Elise; Langelier, Bénédicte; Aïoun, Josiane; Al-Gubory, Kaïs H; Bordat, Christian; Lavialle, Monique; Heberden, Christine

    2016-03-01

    Fatty acid binding protein 7 (FABP7), abundant in the embryonic brain, binds with the highest affinity to docosahexaenoic acid (DHA) and is expressed in the early stages of embryogenesis. Here, we have examined the consequences of the exposure to different DHA levels and of the in utero depletion of FABP7 on early rat brain development. Neurodevelopment was evaluated through the contents of two proteins, connexin 43 (Cx43) and cyclin-dependent kinase 5 (CDK5), both involved in neuroblast proliferation, differentiation, and migration. The dams were fed with diets presenting different DHA contents, from deficiency to supplementation. DHA brain embryos contents already differed at embryonic day 11.5 and the differences kept increasing with time. Cx43 and CDK5 contents were positively associated with the brain DHA levels. When FABP7 was depleted in vivo by injections of siRNA in the telencephalon, the enhancement of the contents of both proteins was lost in supplemented animals, but FABP7 depletion did not modify phospholipid compositions regardless of the diets. Thus, FABP7 is a necessary mediator of the effect of DHA on these proteins synthesis, but its role in DHA uptake is not critical, although FABP7 is localized in phospholipid-rich areas. Our study shows that high contents of DHA associated with FABP7 are necessary to promote early brain development, which prompted us to recommend DHA supplementation early in pregnancy.

  6. Structure of the ordered hydration of amino acids in proteins: analysis of crystal structures

    SciTech Connect

    Biedermannová, Lada Schneider, Bohdan

    2015-10-27

    The hydration of protein crystal structures was studied at the level of individual amino acids. The dependence of the number of water molecules and their preferred spatial localization on various parameters, such as solvent accessibility, secondary structure and side-chain conformation, was determined. Crystallography provides unique information about the arrangement of water molecules near protein surfaces. Using a nonredundant set of 2818 protein crystal structures with a resolution of better than 1.8 Å, the extent and structure of the hydration shell of all 20 standard amino-acid residues were analyzed as function of the residue conformation, secondary structure and solvent accessibility. The results show how hydration depends on the amino-acid conformation and the environment in which it occurs. After conformational clustering of individual residues, the density distribution of water molecules was compiled and the preferred hydration sites were determined as maxima in the pseudo-electron-density representation of water distributions. Many hydration sites interact with both main-chain and side-chain amino-acid atoms, and several occurrences of hydration sites with less canonical contacts, such as carbon–donor hydrogen bonds, OH–π interactions and off-plane interactions with aromatic heteroatoms, are also reported. Information about the location and relative importance of the empirically determined preferred hydration sites in proteins has applications in improving the current methods of hydration-site prediction in molecular replacement, ab initio protein structure prediction and the set-up of molecular-dynamics simulations.

  7. Multi-level machine learning prediction of protein-protein interactions in Saccharomyces cerevisiae.

    PubMed

    Zubek, Julian; Tatjewski, Marcin; Boniecki, Adam; Mnich, Maciej; Basu, Subhadip; Plewczynski, Dariusz

    2015-01-01

    Accurate identification of protein-protein interactions (PPI) is the key step in understanding proteins' biological functions, which are typically context-dependent. Many existing PPI predictors rely on aggregated features from protein sequences, however only a few methods exploit local information about specific residue contacts. In this work we present a two-stage machine learning approach for prediction of protein-protein interactions. We start with the carefully filtered data on protein complexes available for Saccharomyces cerevisiae in the Protein Data Bank (PDB) database. First, we build linear descriptions of interacting and non-interacting sequence segment pairs based on their inter-residue distances. Secondly, we train machine learning classifiers to predict binary segment interactions for any two short sequence fragments. The final prediction of the protein-protein interaction is done using the 2D matrix representation of all-against-all possible interacting sequence segments of both analysed proteins. The level-I predictor achieves 0.88 AUC for micro-scale, i.e., residue-level prediction. The level-II predictor improves the results further by a more complex learning paradigm. We perform 30-fold macro-scale, i.e., protein-level cross-validation experiment. The level-II predictor using PSIPRED-predicted secondary structure reaches 0.70 precision, 0.68 recall, and 0.70 AUC, whereas other popular methods provide results below 0.6 threshold (recall, precision, AUC). Our results demonstrate that multi-scale sequence features aggregation procedure is able to improve the machine learning results by more than 10% as compared to other sequence representations. Prepared datasets and source code for our experimental pipeline are freely available for download from: http://zubekj.github.io/mlppi/ (open source Python implementation, OS independent).

  8. Multi-level machine learning prediction of protein-protein interactions in Saccharomyces cerevisiae.

    PubMed

    Zubek, Julian; Tatjewski, Marcin; Boniecki, Adam; Mnich, Maciej; Basu, Subhadip; Plewczynski, Dariusz

    2015-01-01

    Accurate identification of protein-protein interactions (PPI) is the key step in understanding proteins' biological functions, which are typically context-dependent. Many existing PPI predictors rely on aggregated features from protein sequences, however only a few methods exploit local information about specific residue contacts. In this work we present a two-stage machine learning approach for prediction of protein-protein interactions. We start with the carefully filtered data on protein complexes available for Saccharomyces cerevisiae in the Protein Data Bank (PDB) database. First, we build linear descriptions of interacting and non-interacting sequence segment pairs based on their inter-residue distances. Secondly, we train machine learning classifiers to predict binary segment interactions for any two short sequence fragments. The final prediction of the protein-protein interaction is done using the 2D matrix representation of all-against-all possible interacting sequence segments of both analysed proteins. The level-I predictor achieves 0.88 AUC for micro-scale, i.e., residue-level prediction. The level-II predictor improves the results further by a more complex learning paradigm. We perform 30-fold macro-scale, i.e., protein-level cross-validation experiment. The level-II predictor using PSIPRED-predicted secondary structure reaches 0.70 precision, 0.68 recall, and 0.70 AUC, whereas other popular methods provide results below 0.6 threshold (recall, precision, AUC). Our results demonstrate that multi-scale sequence features aggregation procedure is able to improve the machine learning results by more than 10% as compared to other sequence representations. Prepared datasets and source code for our experimental pipeline are freely available for download from: http://zubekj.github.io/mlppi/ (open source Python implementation, OS independent). PMID:26157620

  9. Site-specific incorporation of keto amino acids into functional G protein-coupled receptors using unnatural amino acid mutagenesis.

    PubMed

    Ye, Shixin; Köhrer, Caroline; Huber, Thomas; Kazmi, Manija; Sachdev, Pallavi; Yan, Elsa C Y; Bhagat, Aditi; RajBhandary, Uttam L; Sakmar, Thomas P

    2008-01-18

    G protein-coupled receptors (GPCRs) are ubiquitous heptahelical transmembrane proteins involved in a wide variety of signaling pathways. The work described here on application of unnatural amino acid mutagenesis to two GPCRs, the chemokine receptor CCR5 (a major co-receptor for the human immunodeficiency virus) and rhodopsin (the visual photoreceptor), adds a new dimension to studies of GPCRs. We incorporated the unnatural amino acids p-acetyl-L-phenylalanine (Acp) and p-benzoyl-L-phenylalanine (Bzp) into CCR5 at high efficiency in mammalian cells to produce functional receptors harboring reactive keto groups at three specific positions. We obtained functional mutant CCR5, at levels up to approximately 50% of wild type as judged by immunoblotting, cell surface expression, and ligand-dependent calcium flux. Rhodopsin containing Acp at three different sites was also purified in high yield (0.5-2 microg/10(7) cells) and reacted with fluorescein hydrazide in vitro to produce fluorescently labeled rhodopsin. The incorporation of reactive keto groups such as Acp or Bzp into GPCRs allows their reaction with different reagents to introduce a variety of spectroscopic and other probes. Bzp also provides the possibility of photo-cross-linking to identify precise sites of protein-protein interactions, including GPCR binding to G proteins and arrestins, and for understanding the molecular basis of ligand recognition by chemokine receptors. PMID:17993461

  10. Trapping the dynamic acyl carrier protein in fatty acid biosynthesis

    PubMed Central

    Nguyen, Chi; Haushalter, Robert W.; Lee, D. John; Markwick, Phineus R. L.; Bruegger, Joel; Caldara-Festin, Grace; Finzel, Kara; Jackson, David R.; Ishikawa, Fumihiro; O’Dowd, Bing; McCammon, J. Andrew; Opella, Stanley J.; Tsai, Shiou-Chuan; Burkart, Michael D.

    2015-01-01

    Acyl carrier protein (ACP) transports the growing fatty acid chain between enzyme domains of fatty acid synthase (FAS) during biosynthesis.1 Because FAS enzymes operate upon ACP-bound acyl groups, ACP must stabilize and transport the growing lipid chain.2 The transient nature of ACP-enzyme interactions imposes a major obstacle to gaining high-resolution structural information about fatty acid biosynthesis, and a new strategy is required to properly study protein-protein interactions. In this work, we describe the application of a mechanism-based probe that allows site-selective covalent crosslinking of AcpP to FabA, the E. coli ACP and fatty acid 3-hydroxyacyl-ACP dehydratase. We report the 1.9 Å crystal structure of the crosslinked AcpP=FabA complex as a homo-dimer, in which AcpP exhibits two different conformations likely representing snapshots of ACP in action: the 4′-phosphopantetheine (PPant) group of AcpP first binds an arginine-rich groove of FabA, followed by an AcpP helical conformational change that locks the AcpP and FabA in place. Residues at the interface of AcpP and FabA are identified and validated by solution NMR techniques, including chemical shift perturbations and RDC measurements. These not only support our interpretation of the crystal structures but also provide an animated view of ACP in action during fatty acid dehydration. Combined with molecular dynamics simulations, we show for the first time that FabA extrudes the sequestered acyl chain from the ACP binding pocket before dehydration by repositioning helix III. Extensive sequence conservation among carrier proteins suggests that the mechanistic insights gleaned from our studies will prove general for fatty acid, polyketide and non-ribosomal biosyntheses. Here the foundation is laid for defining the dynamic action of carrier protein activity in primary and secondary metabolism, providing insight into pathways that can play major roles in the treatment of cancer, obesity and infectious

  11. A comparative study of ribosomal proteins: linkage between amino acid distribution and ribosomal assembly

    PubMed Central

    2013-01-01

    Background Assembly of the ribosome from its protein and RNA constituents must occur quickly and efficiently in order to synthesize the proteins necessary for all cellular activity. Since the early 1960’s, certain characteristics of possible assembly pathways have been elucidated, yet the mechanisms that govern the precise recognition events remain unclear. We utilize a comparative analysis to investigate the amino acid composition of ribosomal proteins (r-proteins) with respect to their role in the assembly process. We compared small subunit (30S) r-protein sequences to those of other housekeeping proteins from 560 bacterial species and searched for correlations between r-protein amino acid content and factors such as assembly binding order, environmental growth temperature, protein size, and contact with ribosomal RNA (rRNA) in the 30S complex. Results We find r-proteins have a significantly high percent of positive residues, which are highly represented at rRNA contact sites. An inverse correlation between the percent of positive residues and r-protein size was identified and is mainly due to the content of Lysine residues, rather than Arginine. Nearly all r-proteins carry a net positive charge, but no statistical correlation between the net charge and the binding order was detected. Thermophilic (high-temperature) r-proteins contain increased Arginine, Isoleucine, and Tyrosine, and decreased Serine and Threonine compared to mesophilic (lower-temperature), reflecting a known distinction between thermophiles and mesophiles, possibly to account for protein thermostability. However, this difference in amino acid content does not extend to rRNA contact sites, as the proportions of thermophilic and mesophilic contact residues are not significantly different. Conclusions Given the significantly higher level of positively charged residues in r-proteins and at contact sites, we conclude that ribosome assembly relies heavily on an electrostatic component of interaction

  12. An amino acid depleted cell-free protein synthesis system for the incorporation of non-canonical amino acid analogs into proteins.

    PubMed

    Singh-Blom, Amrita; Hughes, Randall A; Ellington, Andrew D

    2014-05-20

    Residue-specific incorporation of non-canonical amino acids into proteins is usually performed in vivo using amino acid auxotrophic strains and replacing the natural amino acid with an unnatural amino acid analog. Herein, we present an efficient amino acid depleted cell-free protein synthesis system that can be used to study residue-specific replacement of a natural amino acid by an unnatural amino acid analog. This system combines a simple methodology and high protein expression titers with a high-efficiency analog substitution into a target protein. To demonstrate the productivity and efficacy of a cell-free synthesis system for residue-specific incorporation of unnatural amino acids in vitro, we use this system to show that 5-fluorotryptophan and 6-fluorotryptophan substituted streptavidin retain the ability to bind biotin despite protein-wide replacement of a natural amino acid for the amino acid analog. We envisage this amino acid depleted cell-free synthesis system being an economical and convenient format for the high-throughput screening of a myriad of amino acid analogs with a variety of protein targets for the study and functional characterization of proteins substituted with unnatural amino acids when compared to the currently employed in vivo methodologies.

  13. Effects of Salvianolic Acid B on Protein Expression in Human Umbilical Vein Endothelial Cells

    PubMed Central

    Chang, Tsong-Min; Shi, Guey-Yueh; Wu, Hua-Lin; Wu, Chieh-Hsi; Su, Yan-Di; Wang, Hui-Lin; Wen, Hsin-Yun; Huang, Huey-Chun

    2011-01-01

    Salvianolic acid B (Sal B), a pure water-soluble compound extracted from Radix Salviae miltiorrhizae, has been reported to possess potential cardioprotective efficacy. To identify proteins or pathways by which Sal B might exert its protective activities on the cardiovascular system, two-dimensional gel electrophoresis-based comparative proteomics was performed, and proteins altered in their expression level after Sal B treatment were identified by MALDI-TOF MS/MS. Human umbilical vein endothelial cells (HUVECs) were incubated at Sal B concentrations that can be reached in human plasma by pharmacological intervention. Results indicated that caldesmon, an actin-stabilizing protein, was downregulated in Sal B-exposed HUVECs. Proteins that showed increased expression levels upon Sal B treatment were vimentin, T-complex protein 1, protein disulfide isomerase, tropomyosin alpha, heat shock protein beta-1, UBX domain-containing protein 1, alpha enolase, and peroxiredoxin-2. Additionally, Sal B leads to increased phosphorylation of nucleophosmin in a dose-dependent manner and promotes proliferation of HUVECs. We found that Sal B exhibited a coordinated regulation of enzymes and proteins involved in cytoskeletal reorganization, oxidative stress, and cell growth. Our investigation would provide understanding to the endothelium protection information of Sal B. PMID:21423689

  14. Fatty acid hydroperoxide lyase is a heme protein.

    PubMed

    Shibata, Y; Matsui, K; Kajiwara, T; Hatanaka, A

    1995-02-01

    Fatty acid hydroperoxide lyase (HPO lyase) is an enzyme that cleaves hydroperoxides of polyunsaturated fatty acids to form short chain aldehydes and omega-oxoacids. Spectrophotometric analyses of HPO lyase highly purified from green bell pepper fruits indicate that it is a heme protein. The heme species was revealed to be heme b (protoheme IX) from the absorption spectrum of the pyridine hemochromogen. Although the spectrum highly resembles that of a plant cytochrome P450, allene oxide synthase from flaxseed, CO treatment of the enzyme caused no appearance of a peak at 450 nm, which is an essential diagnostic feature of a cytochrome P450. Internal amino acid sequences determined with peptide fragments obtained from the lyase showed no homology with any reported sequences.

  15. Fatty acids exacerbate tubulointerstitial injury in protein-overload proteinuria.

    PubMed

    Thomas, Mark E; Harris, Kevin P G; Walls, John; Furness, Peter N; Brunskill, Nigel J

    2002-10-01

    The role of the albumin-carried fatty acids in the induction of tubulointerstitial injury was studied in protein-overload proteinuria. Rats were injected with fatty acid-carrying BSA [FA(+)BSA], fatty acid-depleted BSA [FA(-)BSA], or saline. Macrophage infiltration was measured by immunohistochemical staining, apoptotic cells were detected by in situ end labeling, and proliferating cells were identified by in situ hybridization for histone mRNA. Macrophage infiltration was significantly greater in the FA(+)BSA group than in the FA(-)BSA and saline groups. The infiltrate was largely restricted to the outer cortex. Apoptosis was greater in the FA(+)BSA group than in the FA(-)BSA and saline groups. Compared with the saline group, apoptosis was significantly increased in the FA(+)BSA group but not in the FA(-)BSA group. Cortical cells proliferated significantly more in the FA(+)BSA and FA(-)BSA groups than in the saline group. FA(+)BSA is therefore a more potent inducer of macrophage infiltration and cell death than FA(-)BSA. The fatty acids carried on albumin may be the chief instigators of tubulointerstitial injury in protein-overload proteinuria. PMID:12217854

  16. Exploring Sequence Characteristics Related to High-Level Production of Secreted Proteins in Aspergillus niger

    PubMed Central

    van den Berg, Bastiaan A.; Reinders, Marcel J. T.; Hulsman, Marc; Wu, Liang; Pel, Herman J.; Roubos, Johannes A.; de Ridder, Dick

    2012-01-01

    Protein sequence features are explored in relation to the production of over-expressed extracellular proteins by fungi. Knowledge on features influencing protein production and secretion could be employed to improve enzyme production levels in industrial bioprocesses via protein engineering. A large set, over 600 homologous and nearly 2,000 heterologous fungal genes, were overexpressed in Aspergillus niger using a standardized expression cassette and scored for high versus no production. Subsequently, sequence-based machine learning techniques were applied for identifying relevant DNA and protein sequence features. The amino-acid composition of the protein sequence was found to be most predictive and interpretation revealed that, for both homologous and heterologous gene expression, the same features are important: tyrosine and asparagine composition was found to have a positive correlation with high-level production, whereas for unsuccessful production, contributions were found for methionine and lysine composition. The predictor is available online at http://bioinformatics.tudelft.nl/hipsec. Subsequent work aims at validating these findings by protein engineering as a method for increasing expression levels per gene copy. PMID:23049690

  17. Effect of amino acid supplementation on protein quality of soy-based infant formulas fed to rats.

    PubMed

    Sarwar, G; Peace, R W; Botting, H G

    1993-05-01

    The powder forms of soy-based infant formulas obtained from four manufacturers were fed to weanling rats for two weeks, as the sole source of protein in diets containing 8% protein, 20% fat, and adequate amounts of minerals and vitamins. The relative protein efficiency ratio (RPER) and the relative net protein ratio (RNPR) values (casein + methionine = 100) of diets containing unsupplemented formulas were 71-81 and 78-85, respectively. Supplementation of the formula diets with lysine (0.2%), methionine (0.2%), threonine (0.1%) or tryptophan (0.05%) increased the level of the supplemental amino acid in rat serum but generally failed to improve the RPER or RNPR values. Addition of all four essential amino acids to the formula diets, however, caused a marked improvement in their protein quality (RPER or RNPR values = 100). The data suggested that proteins in soy-based formulas could be marginally co-limited in several indispensable amino acids.

  18. Hydrolysed proteins in preterm formula: influence on plasma aminoacids, blood fatty acids and insulinaemia.

    PubMed

    Agosti, M; Pugni, L; Ramenghi, L A; Mosca, F; Marini, A

    2003-09-01

    The data from the present investigation differ from those of the previous study. The new version of hydrolysed protein formula did not induce changes in insulinaemia or in the insulinaemia/glycaemia ratio in pre- and postprandial samples when compared with the intact protein formula. The investigation also confirmed that branch chain aminoacids regulate insulin secretion and that the length of chain is insulinotropic for fatty acids, while the degree of unsaturation is not able to reduce insulin secretion in the newborn baby. This could be caused by stimulation of insulin output activated by the C20:5 n 3; the effects of fatty acids are probably related more to the variable composition of cell membrane than to the fatty acids circulating levels.

  19. Plant growth, metabolism and adaptation in relation to stress conditions. XXVII. Can ascorbic acid modify the adverse effects of NaCl and mannitol on amino acids, nucleic acids and protein patterns in Vicia faba seedlings?

    PubMed

    Younis, M E; Hasaneen, M N A; Kazamel, A M S

    2009-03-01

    The adverse effects of either NaCl or mannitol on amino acids, protein patterns and nucleic acids in Vicia faba seeds were investigated. The exogenous addition of 4 mM ascorbic acid to the stressing media in which the broad bean seeds were germinated in combination with either the ionic (NaCl) or osmotic (mannitol) stressor induced significant protective changes in the total amount and in the relative composition of amino acids in general and in proline, glycine, glutamic, aspartic, alanine and serine in particular. It also induced changes in nucleic acids (RNA and DNA) content. These changes occurred throughout the entire period of the experiments (12 days). Separate administration of NaCl or mannitol enhanced the occurrence of particular novel proteins that were not detected in control bean seeds (water medium). Protein banding patterns of broad bean seedlings treated with NaCl or mannitol in combination with 4 mM ascorbic acid showed different de novo protein bands, with different molecular weights, at different stages of seedlings growth, with lower levels or a nearly complete absence of the major stress proteins. The pattern of changes for amino acids and nucleic acids and the range of protein bands extracted from the variously treated broad bean seedlings indicate a positive role of ascorbic acid in the alleviation of the damage effects induced by NaCl and mannitol. The importance of this role in the stress tolerance of broad beans is discussed.

  20. Assembly of the bacteriophage T4 replication machine requires the acidic carboxy terminus of gene 32 protein.

    PubMed

    Hurley, J M; Chervitz, S A; Jarvis, T C; Singer, B S; Gold, L

    1993-01-20

    The acidic carboxy-terminal 89-amino acid fragment of bacteriophage T4 gene 32 protein was expressed in Escherichia coli to high levels from an inducible plasmid construct. Infection of induced cells by wild-type T4 phage results in impaired phage DNA synthesis. The time at which DNA synthesis begins and the diminution in DNA synthesis rates correlate with the amount of carboxy-terminal peptide that accumulates intracellularly prior to infection. Correspondingly, when induced cells are infected with viable phage containing a small deletion near the carboxy-terminus of 32 protein (delta PR201), the inhibition of phage DNA synthesis was much more severe. The mutant 32 protein competes less well against overproduced wild-type acid peptide than does wild-type 32 protein. The purified acid peptide, when used as the attached ligand for affinity chromatography, binds several T4 proteins from phage-infected cells, including 43 protein (T4 DNA polymerase), Dda protein (a DNA helicase), and UvsX protein (a Rec-like recombination protein). Furthermore, at 50- to 100-fold molar excess of acid peptide over intact 32 protein, phage DNA synthesis was specifically inhibited at the initiation step in an in vitro 5-protein DNA replication experiment. We propose that one or more phage replication proteins are titrated as non-productive protein-protein complexes at a site away from the DNA template. This implies that the carboxy-terminal domain of 32 protein is involved in an obligate step of replication machine assembly when the protein is properly attached to ssDNA in the vicinity of a primer-template junction. The assembly defect we observe is strikingly similar to the repression, or "squelching", of the activity of certain eukaryotic transcriptional activators. PMID:8429554

  1. Em polypeptide and its messenger RNA levels are modulated by abscisic acid during embryogenesis in wheat.

    PubMed

    Williamson, J D; Quatrano, R S; Cuming, A C

    1985-10-15

    The effect of abscisic acid (ABA) on the expression of the 'early-methionine-labeled' (Em) polypeptide was examined in cultured, immature wheat (Triticum aestivum, L.) embryos and in developing embryos in planta. A complementary DNA (cDNA) library was constructed from poly(A)-rich RNA from immature embryos cultured in the presence of ABA. ABA-enhanced sequences were first identified by differential colony-blot hybridization, and then verified using RNA slot-blot analysis. Dot-blot hybridization showed that one clone, p1015, was homologous to the previously isolated Em cDNA, pWG432. Electrophoretic analysis of the hybrid-select translation product of p1015 confirmed its identity as an Em sequence. Comparison of the p1015 cDNA insert size and the Em message size, from northern blot analysis, showed that p1015 contained about 87% of the Em sequence. RNA slot-blot analysis and protein electrophoresis showed that Em message, but not Em protein, accumulated at a low, basal level in immature embryos in the absence of ABA. Neither Em message nor Em protein was seen in three-day germinated seedlings. Steady-state levels of Em message and protein increased in immature embryos in the presence of ABA, both in culture and in planta. Regulation appeared to be primarily at the level of transcription or specific message stability. Regulation may also involve specific protein stability, since synthesis of Em protein continued in immature embryos in the absence of ABA, but Em protein did not accumulate in detectable amounts. We conclude that ABA specifically modulates Em message and protein levels in immature embryos, but is probably not responsible for the embryogenic specificity of Em expression.

  2. Effect of energy and protein levels on nutrient utilization and their requirements in growing Murrah buffaloes.

    PubMed

    Prusty, Sonali; Kundu, Shivlal Singh; Mondal, Goutam; Sontakke, Umesh; Sharma, Vijay Kumar

    2016-04-01

    To evaluate different levels of energy and protein for optimum growth of Murrah male buffalo calves, a growth trial (150 days) was conducted on 30 calves (body weight 202.5 ± 6.8 kg). Six diets were formulated to provide 90, 100 and 110% protein level and 90 and 110% energy level requirements for buffalo calves, derived from ICAR 2013 recommendations for buffaloes. The crude protein (CP) intake was increased with higher dietary CP, whereas no effect of energy levels or interaction between protein and energy was observed on CP intake. There were significant effects (P < 0.01) of the interaction between protein and energy (P < 0.05) on metabolizable energy (ME) intake. The digestibility of dry matter (DM), organic matter (OM) and non-fibrous carbohydrate (NFC) was higher (P < 0.0001) in high-energy groups compared to low-energy groups. The CP digestibility increased with the increased CP and ME of the rations. The absorbed N was improved linearly with an increased level of dietary CP, whereas the N retention was similar among all the groups distributed as per different energy or protein levels. The nutrient intake (protein or energy) per kg body weight (BW)(0.75) at various fortnight intervals was regressed linearly from the average daily gain (ADG) per kg BW(0.75). By setting the average daily gain at zero in the developed regression equation, a maintenance requirement was obtained, i.e. 133.1 kcal ME, 6.45 g CP and 3.95 g metabolizable protein (MP) per kg BW(0.75). Requirement for growth was 6.12 kcal ME, 0.46 g CP and 0.32 g MP per kg BW(0.75) per day. Metabolizable amino acid requirement was estimated from partitioning of MP intake and ADG. The ME requirements were lower, whereas the MP requirement of Murrah buffaloes was higher than ICAR (2013) recommendations. PMID:26970972

  3. Smoking, COPD and 3-Nitrotyrosine Levels of Plasma Proteins

    SciTech Connect

    Jin, Hongjun; Webb-Robertson, Bobbie-Jo M.; Peterson, Elena S.; Tan, Ruimin; Bigelow, Diana J.; Scholand, Mary Beth; Hoidal, John R.; Pounds, Joel G.; Zangar, Richard C.

    2011-09-01

    BACKGROUND: Nitric oxide is a physiologically regulator of endothelial function and hemodynamics. Oxidized products of nitric oxide can form nitrotyrosine, which is a marker of nitrative stress. Cigarette smoking decreases exhaled nitric oxide, and the underlying mechanism may be important in the cardiovascular toxicity of cigarette smoke, although it is not clear if this effect results from decreased nitric oxide production or oxidation of nitric oxide to reactive, nitrating, species. These processes would be expected to have opposite effects on nitrotyrosine levels, a marker of nitrative stress. OBJECTIVE: In this study, we determine the effects of smoking and chronic obstructive pulmonary disease (COPD) on circulating levels of nitrotyrosine, and thereby gain insight into the processes regulating nitrotyrosine formation. METHODS: A custom antibody microarray platform was used to analyze the levels of 3-nitrotyrosine modifications on 24 proteins in plasma. Plasma samples from 458 individuals were analyzed. RESULTS: Nitrotyrosine levels in circulating proteins were uniformly reduced in smokers but increased in COPD patients. We also observed a persistent suppression of nitrotyrosine in former smokers. CONCLUSIONS: Smoking broadly suppresses the levels of 3-nitrotyrosine in plasma proteins, suggesting that cigarette smoke suppresses endothelial nitric oxide production. In contrast, the increase in nitrotyrosine levels in COPD patients most likely results from inflammatory processes. This study provides the first evidence that smoking has irreversible effects on endothelial production of nitric oxide, and provides insight into how smoking could induce a loss of elasticity in the vasculature and a long-term increase in the risk of cardiovascular disease.

  4. GPCR-MPredictor: multi-level prediction of G protein-coupled receptors using genetic ensemble.

    PubMed

    Naveed, Muhammad; Khan, Asifullah; Khan, Asif Ullah

    2012-05-01

    G protein-coupled receptors (GPCRs) are transmembrane proteins, which transduce signals from extracellular ligands to intracellular G protein. Automatic classification of GPCRs can provide important information for the development of novel drugs in pharmaceutical industry. In this paper, we propose an evolutionary approach, GPCR-MPredictor, which combines individual classifiers for predicting GPCRs. GPCR-MPredictor is a web predictor that can efficiently predict GPCRs at five levels. The first level determines whether a protein sequence is a GPCR or a non-GPCR. If the predicted sequence is a GPCR, then it is further classified into family, subfamily, sub-subfamily, and subtype levels. In this work, our aim is to analyze the discriminative power of different feature extraction and classification strategies in case of GPCRs prediction and then to use an evolutionary ensemble approach for enhanced prediction performance. Features are extracted using amino acid composition, pseudo amino acid composition, and dipeptide composition of protein sequences. Different classification approaches, such as k-nearest neighbor (KNN), support vector machine (SVM), probabilistic neural networks (PNN), J48, Adaboost, and Naives Bayes, have been used to classify GPCRs. The proposed hierarchical GA-based ensemble classifier exploits the prediction results of SVM, KNN, PNN, and J48 at each level. The GA-based ensemble yields an accuracy of 99.75, 92.45, 87.80, 83.57, and 96.17% at the five levels, on the first dataset. We further perform predictions on a dataset consisting of 8,000 GPCRs at the family, subfamily, and sub-subfamily level, and on two other datasets of 365 and 167 GPCRs at the second and fourth levels, respectively. In comparison with the existing methods, the results demonstrate the effectiveness of our proposed GPCR-MPredictor in classifying GPCRs families. It is accessible at http://111.68.99.218/gpcr-mpredictor/.

  5. Direct protein-protein interactions and substrate channeling between cellular retinoic acid binding proteins and CYP26B1.

    PubMed

    Nelson, Cara H; Peng, Chi-Chi; Lutz, Justin D; Yeung, Catherine K; Zelter, Alex; Isoherranen, Nina

    2016-08-01

    Cellular retinoic acid binding proteins (CRABPs) bind all-trans-retinoic acid (atRA) tightly. This study aimed to determine whether atRA is channeled directly to cytochrome P450 (CYP) CYP26B1 by CRABPs, and whether CRABPs interact directly with CYP26B1. atRA bound to CRABPs (holo-CRABP) was efficiently metabolized by CYP26B1. Isotope dilution experiments showed that delivery of atRA to CYP26B1 in solution was similar with or without CRABP. Holo-CRABPs had higher affinity for CYP26B1 than free atRA, but both apo-CRABPs inhibited the formation of 4-OH-RA by CYP26B1. Similar protein-protein interactions between soluble binding proteins and CYPs may be important for other lipophilic CYP substrates.

  6. Serum fluoride and sialic acid levels in osteosarcoma.

    PubMed

    Sandhu, R; Lal, H; Kundu, Z S; Kharb, S

    2011-12-01

    Osteosarcoma is a rare malignant bone tumor most commonly occurring in children and young adults presenting with painful swelling. Various etiological factors for osteosarcoma are ionizing radiation, family history of bone disorders and cancer, chemicals (fluoride, beryllium, and vinyl chloride), and viruses. Status of fluoride levels in serum of osteosarcoma is still not clear. Recent reports have indicated that there is a link between fluoride exposure and osteosarcoma. Glycoproteins and glycosaminoglycans are an integral part of bone and prolonged exposure to fluoride for long duration has been shown to cause degradation of collagen and ground substance in bones. The present study was planned to analyze serum fluoride, sialic acid, calcium, phosphorus, and alkaline phosphatase levels in 25 patients of osteosarcoma and age- and sex-matched subjects with bone-forming tumours other than osteosarcoma and musculo-skeletal pain (controls, 25 each). Fluoride levels were analyzed by ISE and sialic acid was analyzed by Warren's method. Mean serum fluoride concentration was found to be significantly higher in patients with osteosarcoma as compared to the other two groups. The mean value of flouride in patients with other bone-forming tumors was approximately 50% of the group of osteosarcoma; however, it was significantly higher when compared with patients of group I. Serum sialic acid concentration was found to be significantly raised in patients with osteosarcoma as well as in the group with other bone-forming tumors as compared to the group of controls. There was, however, no significant difference in the group of patients of osteosarcoma when compared with group of patients with other bone-forming tumors. These results showing higher level of fluoride with osteosarcoma compared to others suggesting a role of fluoride in the disease.

  7. Prognostic Significance of Uric Acid Levels in Ischemic Stroke Patients.

    PubMed

    Zhang, Xia; Huang, Zhi-Chao; Lu, Tao-Sheng; You, Shou-Jiang; Cao, Yong-Jun; Liu, Chun-Feng

    2016-01-01

    The importance and function of serum uric acid (UA) levels in patients with cardiovascular disease or stroke are unclear. We sought to evaluate the appropriate UA levels for stroke patients and the association between endogenous UA levels and clinical outcomes in acute ischemic stroke (AIS) patients, particularly regarding the possible interaction between gender and UA levels with respect to AIS prognosis. We examined 303 patients who had an onset of ischemic stroke within 48 h. Of those, 101 patients received thrombolytic treatment. Serum UA (μmol/L) levels were measured the second morning after admission. Patient prognosis was evaluated 90 days after clinical onset by modified Rankin Scale. Patients were divided into four groups according to serum UA quartiles. A binary multivariate logistic regression model was used to assess clinical relevance in regard to functional outcome and endogenous UA levels. Analysis of subgroups by gender and normal glomerular filtration rate were also been done. Poor functional outcome was associated with older age, history of atrial fibrillation, or higher baseline National Institutes of Health Stroke Scale scores. After adjustment for potential confounders, patients with higher UA levels (>380 μmol/L) or lower UA levels (≤250 μmol/L) were 2-3 times more likely to have a poor outcome (OR 2.95, 95% CI 1.14-7.61; OR 2.78, 95% CI 1.02-7.58, respectively) compared to the baseline group (UA level 316-380 μmol/L). The same results were observed in thrombolyzed patients. Patients with high and low UA levels were 9-18 times more likely to having poor outcomes compared to the baseline group (UA level: 316-380 μmol/L; OR 18.50, 95% CI: 2.041-167.67; OR 9.66, 95% CI 1.42-65.88, respectively). In men, patients with high UA levels were 6 times more likely to have poor outcomes compared to the baseline group (UA level: 279-334 μmol/L; OR 6.10, 95% CI 1.62-22.93). However, female patients with UA level 271-337 μmol/L were seven times more

  8. Mosaic protein and nucleic acid vaccines against hepatitis C virus

    SciTech Connect

    Yusim, Karina; Korber, Bette T. M.; Kuiken, Carla L.; Fischer, William M.

    2013-06-11

    The invention relates to immunogenic compositions useful as HCV vaccines. Provided are HCV mosaic polypeptide and nucleic acid compositions which provide higher levels of T-cell epitope coverage while minimizing the occurrence of unnatural and rare epitopes compared to natural HCV polypeptides and consensus HCV sequences.

  9. Cystine levels, cystine flux, and protein catabolism in cancer cachexia, HIV/SIV infection, and senescence.

    PubMed

    Hack, V; Schmid, D; Breitkreutz, R; Stahl-Henning, C; Drings, P; Kinscherf, R; Taut, F; Holm, E; Dröge, W

    1997-01-01

    Patients with skeletal muscle catabolism (cachexia) fail to conserve the skeletal muscle protein and release large amounts of nitrogen as urea. Previous studies suggest that the threshold for the conversion of amino acids into other forms of chemical energy and the concomitant production of urea are regulated by the plasma cystine level and hepatic cysteine catabolism. Studies of plasma amino acid exchange rates in the lower extremities now show that healthy young subjects regulate their plasma cystine level in a process that may be described as controlled constructive catabolism. The term controlled describes the fact that the release of cystine and other amino acids from the peripheral tissue is negatively correlated with (certain) plasma amino acid levels. The term constructive describes the fact that the release of cystine is correlated with an increase of the plasma cystine level. The regulation of the plasma cystine level is disturbed in conditions with progressive skeletal muscle catabolism including cancer, HIV infection, and old age. These conditions show also a low plasma glutamine:cystine ratio indicative of an impaired hepatic cystine catabolism. In HIV+ patients and SIV-infected macaques, a decrease of the plasma cystine level was found to coincide with the decrease of CD4+ T cells.

  10. Characterization of a fatty acid-binding protein from rat heart.

    PubMed

    Offner, G D; Troxler, R F; Brecher, P

    1986-04-25

    A fatty acid-binding protein has been isolated from rat heart and purified by gel filtration chromatography on Sephadex G-75 and anion-exchange chromatography on DE52. The circular dichroic spectrum of this protein was not affected by protein concentration, suggesting that it does not aggregate into multimers. Computer analyses of the circular dichroic spectrum predicted that rat heart fatty acid-binding protein contains approximately 22% alpha-helix, 45% beta-form and 33% unordered structure. Immunological studies showed that the fatty acid-binding proteins from rat heart and rat liver are immunochemically unrelated. The amino acid composition and partial amino acid sequence of the heart protein indicated that it is structurally related to, but distinct from, other fatty acid-binding proteins from liver, intestine, and 3T3 adipocytes. Using a binding assay which measures the transfer of fatty acids between donor liposomes and protein (Brecher, P., Saouaf, R., Sugarman, J. M., Eisenberg, D., and LaRosa, K. (1984) J. Biol. Chem. 259, 13395-13401), it was shown that both rat heart and liver fatty acid-binding proteins bind 2 mol of oleic acid or palmitic acid/mol of protein. The structural and functional relationship of rat heart fatty acid-binding protein to fatty acid-binding proteins from other tissues is discussed. PMID:3957934

  11. Post-exercise whey protein hydrolysate supplementation induces a greater increase in muscle protein synthesis than its constituent amino acid content.

    PubMed

    Kanda, Atsushi; Nakayama, Kyosuke; Fukasawa, Tomoyuki; Koga, Jinichiro; Kanegae, Minoru; Kawanaka, Kentaro; Higuchi, Mitsuru

    2013-09-28

    It is well known that ingestion of a protein source is effective in stimulating muscle protein synthesis after exercise. In addition, there are numerous reports on the impact of leucine and leucine-rich whey protein on muscle protein synthesis and mammalian target of rapamycin (mTOR) signalling. However, there is only limited information on the effects of whey protein hydrolysates (WPH) on muscle protein synthesis and mTOR signalling. The aim of the present study was to compare the effects of WPH and amino acids on muscle protein synthesis and the initiation of translation in skeletal muscle during the post-exercise phase. Male Sprague–Dawley rats swam for 2 h to depress muscle protein synthesis. Immediately after exercise, the animals were administered either carbohydrate (CHO), CHO plus an amino acid mixture (AA) or CHO plus WPH. At 1 h after exercise, the supplements containing whey-based protein (AA and WPH) caused a significant increase in the fractional rate of protein synthesis (FSR) compared with CHO. WPH also caused a significant increase in FSR compared with AA. Post-exercise ingestion of WPH caused a significant increase in the phosphorylation of mTOR levels compared with AA or CHO. In addition, WPH caused greater phosphorylation of ribosomal protein S6 kinase and eukaryotic initiation factor 4E-binding protein 1 than AA and CHO. In contrast, there was no difference in plasma amino acid levels following supplementation with either AA or WPH. These results indicate that WPH may include active components that are superior to amino acids for stimulating muscle protein synthesis and initiating translation.

  12. Light-Induced Alterations in Basil Ganglia Kynurenic Acid Levels

    NASA Technical Reports Server (NTRS)

    Sroufe, Angela E.; Whittaker, J. A.; Patrickson, J. W.; Orr, M. C.

    1997-01-01

    The metabolic synthesis, release and breakdown of several known CNS neurotransmitters have been shown to follow a circadian pattern entrained to the environmental light/dark cycle. The levels of excitatory amino acid (EAA) transmitters such as glutamate, have been shown to vary with environmental lighting conditions. Kynurenic Acid (KA), an endogenous tryptophan metabolite and glutamate receptor antagonist, has been reported to have neuroprotective effects against EAA-induced excitotoxic cell damage. Changes in KA's activity within the mammalian basal ganglia has been proposed as being contributory to neurotoxicity in Huntington's Disease. It is not known whether CNS KA levels follow a circadian pattern or exhibit light-induced fluctuations. However, because the symptoms of certain degenerative motor disorders seem to fluctuate with daily 24 hour rhythm, we initiated studies to determine if basal ganglia KA were influenced by the daily light/dark cycle and could influence motor function. Therefore in this study, HPLC-EC was utilized to determine if basal ganglia KA levels in tissue extracts from adult male Long-Evans rats (200-250g) entrained to 24 and 48 hours constant light and dark conditions, respectively. Samples were taken one hour before the onset of the subjective day and one hour prior to the onset of the subjective night in order to detect possible phase differences in KA levels and to allow for accumulation of factors expressed in association with the light or dark phase. Data analysis revealed that KA levels in the basal ganglia vary with environmental lighting conditions; being elevated generally during the dark. Circadian phase differences in KA levels were also evident during the subjective night and subjective day, respectively. Results from these studies are discussed with respect to potential cyclic changes in neuronal susceptibility to excitotoxic damage during the daily 24 hour cycle and its possible relevance to future therapeutic approaches in

  13. The fusogenic lipid phosphatidic acid promotes the biogenesis of mitochondrial outer membrane protein Ugo1

    PubMed Central

    Keller, Michael; Taskin, Asli A.; Horvath, Susanne E.; Guan, Xue Li; Prinz, Claudia; Opalińska, Magdalena; Zorzin, Carina; van der Laan, Martin; Wenk, Markus R.; Schubert, Rolf; Wiedemann, Nils; Holzer, Martin

    2015-01-01

    Import and assembly of mitochondrial proteins depend on a complex interplay of proteinaceous translocation machineries. The role of lipids in this process has been studied only marginally and so far no direct role for a specific lipid in mitochondrial protein biogenesis has been shown. Here we analyzed a potential role of phosphatidic acid (PA) in biogenesis of mitochondrial proteins in Saccharomyces cerevisiae. In vivo remodeling of the mitochondrial lipid composition by lithocholic acid treatment or by ablation of the lipid transport protein Ups1, both leading to an increase of mitochondrial PA levels, specifically stimulated the biogenesis of the outer membrane protein Ugo1, a component of the mitochondrial fusion machinery. We reconstituted the import and assembly pathway of Ugo1 in protein-free liposomes, mimicking the outer membrane phospholipid composition, and found a direct dependency of Ugo1 biogenesis on PA. Thus, PA represents the first lipid that is directly involved in the biogenesis pathway of a mitochondrial membrane protein. PMID:26347140

  14. RNA-binding protein QKI regulates Glial fibrillary acidic protein expression in human astrocytes.

    PubMed

    Radomska, Katarzyna J; Halvardson, Jonatan; Reinius, Björn; Lindholm Carlström, Eva; Emilsson, Lina; Feuk, Lars; Jazin, Elena

    2013-04-01

    Linkage, association and expression studies previously pointed to the human QKI, KH domain containing, RNA-binding (QKI) as a candidate gene for schizophrenia. Functional studies of the mouse orthologue Qk focused mainly on its role in oligodendrocyte development and myelination, while its function in astroglia remained unexplored. Here, we show that QKI is highly expressed in human primary astrocytes and that its splice forms encode proteins targeting different subcellular localizations. Uncovering the role of QKI in astrocytes is of interest in light of growing evidence implicating astrocyte dysfunction in the pathogenesis of several disorders of the central nervous system. We selectively silenced QKI splice variants in human primary astrocytes and used RNA sequencing to identify differential expression and splice variant composition at the genome-wide level. We found that an mRNA expression of Glial fibrillary acidic protein (GFAP), encoding a major component of astrocyte intermediate filaments, was down-regulated after QKI7 splice variant silencing. Moreover, we identified a potential QKI-binding site within the 3' untranslated region of human GFAP. This sequence was not conserved between mice and humans, raising the possibility that GFAP is a target for QKI in humans but not rodents. Haloperidol treatment of primary astrocytes resulted in coordinated increases in QKI7 and GFAP expression. Taken together, our results provide the first link between QKI and GFAP, two genes with alterations previously observed independently in schizophrenic patients. Our findings for QKI, together with its well-known role in myelination, suggest that QKI is a hub regulator of glia function in humans.

  15. Formation, reactivity and detection of protein sulfenic acids

    PubMed Central

    Kettenhofen, Nicholas J.; Wood, Matthew J.

    2010-01-01

    It has become clear in recent decades that the post-translational modification of protein cysteine residues is a crucial regulatory event in biology. Evidence supports the reversible oxidation of cysteine thiol groups as a mechanism of redox-based signal transduction while the accumulation of proteins with irreversible thiol oxidations is a hallmark of stress-induced cellular damage. The initial formation of cysteine sulfenic acid (SOH) derivatives, along with the reactive properties of this functional group, serves as a crossroads whereby the local redox environment may dictate the progression of either regulatory or pathological outcomes. Protein-SOH are established as transient intermediates in the formation of more stable cysteine oxidation products both under basal conditions and in response to several redox-active extrinsic compounds. This review details both direct and multi-step chemical routes proposed to generate protein-SOH, the spectrum of secondary reactions that may follow their initial formation and the arsenal of experimental tools available for their detection. Both the pioneering studies that have provided a framework for our current understanding of protein-SOH as well as state-of-the-art proteomic strategies designed for global assessments of this post-translational modification are highlighted. PMID:20845928

  16. CMP-N-acetylneuraminic acid synthetase interacts with fragile X related protein 1

    PubMed Central

    Ma, Yun; Tian, Shuai; Wang, Zongbao; Wang, Changbo; Chen, Xiaowei; Li, Wei; Yang, Yang; He, Shuya

    2016-01-01

    Fragile X mental retardation protein (FMRP), fragile X related 1 protein (FXR1P) and FXR2P are the members of the FMR protein family. These proteins contain two KH domains and a RGG box, which are characteristic of RNA binding proteins. The absence of FMRP, causes fragile X syndrome (FXS), the leading cause of hereditary mental retardation. FXR1P is expressed throughout the body and important for normal muscle development, and its absence causes cardiac abnormality. To investigate the functions of FXR1P, a screen was performed to identify FXR1P-interacting proteins and determine the biological effect of the interaction. The current study identified CMP-N-acetylneuraminic acid synthetase (CMAS) as an interacting protein using the yeast two-hybrid system, and the interaction between FXR1P and CMAS was validated in yeast using a β-galactosidase assay and growth studies with selective media. Furthermore, co-immunoprecipitation was used to analyze the FXR1P/CMAS association and immunofluorescence microscopy was performed to detect expression and intracellular localization of the proteins. The results of the current study indicated that FXR1P and CMAS interact, and colocalize in the cytoplasm and the nucleus of HEK293T and HeLa cells. Accordingly, a fragile X related 1 (FXR1) gene overexpression vector was constructed to investigate the effect of FXR1 overexpression on the level of monosialotetrahexosylganglioside 1 (GM1). The results of the current study suggested that FXR1P is a tissue-specific regulator of GM1 levels in SH-SY5Y cells, but not in HEK293T cells. Taken together, the results initially indicate that FXR1P interacts with CMAS, and that FXR1P may enhance the activation of sialic acid via interaction with CMAS, and increase GM1 levels to affect the development of the nervous system, thus providing evidence for further research into the pathogenesis of FXS. PMID:27357083

  17. Predicting Protein-Protein Interactions from the Molecular to the Proteome Level.

    PubMed

    Keskin, Ozlem; Tuncbag, Nurcan; Gursoy, Attila

    2016-04-27

    Identification of protein-protein interactions (PPIs) is at the center of molecular biology considering the unquestionable role of proteins in cells. Combinatorial interactions result in a repertoire of multiple functions; hence, knowledge of PPI and binding regions naturally serve to functional proteomics and drug discovery. Given experimental limitations to find all interactions in a proteome, computational prediction/modeling of protein interactions is a prerequisite to proceed on the way to complete interactions at the proteome level. This review aims to provide a background on PPIs and their types. Computational methods for PPI predictions can use a variety of biological data including sequence-, evolution-, expression-, and structure-based data. Physical and statistical modeling are commonly used to integrate these data and infer PPI predictions. We review and list the state-of-the-art methods, servers, databases, and tools for protein-protein interaction prediction. PMID:27074302

  18. Foamy Virus Protein-Nucleic Acid Interactions during Particle Morphogenesis.

    PubMed

    Hamann, Martin V; Lindemann, Dirk

    2016-01-01

    Compared with orthoretroviruses, our understanding of the molecular and cellular replication mechanism of foamy viruses (FVs), a subfamily of retroviruses, is less advanced. The FV replication cycle differs in several key aspects from orthoretroviruses, which leaves established retroviral models debatable for FVs. Here, we review the general aspect of the FV protein-nucleic acid interactions during virus morphogenesis. We provide a summary of the current knowledge of the FV genome structure and essential sequence motifs required for RNA encapsidation as well as Gag and Pol binding in combination with details about the Gag and Pol biosynthesis. This leads us to address open questions in FV RNA engagement, binding and packaging. Based on recent findings, we propose to shift the point of view from individual glycine-arginine-rich motifs having functions in RNA interactions towards envisioning the FV Gag C-terminus as a general RNA binding protein module. We encourage further investigating a potential new retroviral RNA packaging mechanism, which seems more complex in terms of the components that need to be gathered to form an infectious particle. Additional molecular insights into retroviral protein-nucleic acid interactions help us to develop safer, more specific and more efficient vectors in an era of booming genome engineering and gene therapy approaches. PMID:27589786

  19. Lithium Decreases Glial Fibrillary Acidic Protein in a Mouse Model of Alexander Disease.

    PubMed

    LaPash Daniels, Christine M; Paffenroth, Elizabeth; Austin, Elizabeth V; Glebov, Konstantin; Lewis, Diana; Walter, Jochen; Messing, Albee

    2015-01-01

    Alexander disease is a fatal neurodegenerative disease caused by mutations in the astrocyte intermediate filament glial fibrillary acidic protein (GFAP). The disease is characterized by elevated levels of GFAP and the formation of protein aggregates, known as Rosenthal fibers, within astrocytes. Lithium has previously been shown to decrease protein aggregates by increasing the autophagy pathway for protein degradation. In addition, lithium has also been reported to decrease activation of the transcription factor STAT3, which is a regulator of GFAP transcription and astrogliogenesis. Here we tested whether lithium treatment would decrease levels of GFAP in a mouse model of Alexander disease. Mice with the Gfap-R236H point mutation were fed lithium food pellets for 4 to 8 weeks. Four weeks of treatment with LiCl at 0.5% in food pellets decreased GFAP protein and transcripts in several brain regions, although with mild side effects and some mortality. Extending the duration of treatment to 8 weeks resulted in higher mortality, and again with a decrease in GFAP in the surviving animals. Indicators of autophagy, such as LC3, were not increased, suggesting that lithium may decrease levels of GFAP through other pathways. Lithium reduced the levels of phosphorylated STAT3, suggesting this as one pathway mediating the effects on GFAP. In conclusion, lithium has the potential to decrease GFAP levels in Alexander disease, but with a narrow therapeutic window separating efficacy and toxicity.

  20. A simple atomic-level hydrophobicity scale reveals protein interfacial structure.

    PubMed

    Kapcha, Lauren H; Rossky, Peter J

    2014-01-23

    Many amino acid residue hydrophobicity scales have been created in an effort to better understand and rapidly characterize water-protein interactions based only on protein structure and sequence. There is surprisingly low consistency in the ranking of residue hydrophobicity between scales, and their ability to provide insightful characterization varies substantially across subject proteins. All current scales characterize hydrophobicity based on entire amino acid residue units. We introduce a simple binary but atomic-level hydrophobicity scale that allows for the classification of polar and non-polar moieties within single residues, including backbone atoms. This simple scale is first shown to capture the anticipated hydrophobic character for those whole residues that align in classification among most scales. Examination of a set of protein binding interfaces establishes good agreement between residue-based and atomic-level descriptions of hydrophobicity for five residues, while the remaining residues produce discrepancies. We then show that the atomistic scale properly classifies the hydrophobicity of functionally important regions where residue-based scales fail. To illustrate the utility of the new approach, we show that the atomic-level scale rationalizes the hydration of two hydrophobic pockets and the presence of a void in a third pocket within a single protein and that it appropriately classifies all of the functionally important hydrophilic sites within two otherwise hydrophobic pores. We suggest that an atomic level of detail is, in general, necessary for the reliable depiction of hydrophobicity for all protein surfaces. The present formulation can be implemented simply in a manner no more complex than current residue-based approaches.

  1. Differential Acute and Chronic Effects of Leptin on Hypothalamic Astrocyte Morphology and Synaptic Protein Levels

    PubMed Central

    García-Cáceres, Cristina; Fuente-Martín, Esther; Burgos-Ramos, Emma; Granado, Miriam; Frago, Laura M.; Barrios, Vicente; Horvath, Tamas

    2011-01-01

    Astrocytes participate in neuroendocrine functions partially through modulation of synaptic input density in the hypothalamus. Indeed, glial ensheathing of neurons is modified by specific hormones, thus determining the availability of neuronal membrane space for synaptic inputs, with the loss of this plasticity possibly being involved in pathological processes. Leptin modulates synaptic inputs in the hypothalamus, but whether astrocytes participate in this action is unknown. Here we report that astrocyte structural proteins, such as glial fibrillary acidic protein (GFAP) and vimentin, are induced and astrocyte morphology modified by chronic leptin administration (intracerebroventricular, 2 wk), with these changes being inversely related to modifications in synaptic protein densities. Similar changes in glial structural proteins were observed in adult male rats that had increased body weight and circulating leptin levels due to neonatal overnutrition (overnutrition: four pups/litter vs. control: 12 pups/litter). However, acute leptin treatment reduced hypothalamic GFAP levels and induced synaptic protein levels 1 h after administration, with no effect on vimentin. In primary hypothalamic astrocyte cultures leptin also reduced GFAP levels at 1 h, with an induction at 24 h, indicating a possible direct effect of leptin. Hence, one mechanism by which leptin may affect metabolism is by modifying hypothalamic astrocyte morphology, which in turn could alter synaptic inputs to hypothalamic neurons. Furthermore, the responses to acute and chronic leptin exposure are inverse, raising the possibility that increased glial activation in response to chronic leptin exposure could be involved in central leptin resistance. PMID:21343257

  2. Amino acid profiles and digestible indispensable amino acid scores of proteins from the prioritized key foods in Bangladesh.

    PubMed

    Shaheen, Nazma; Islam, Saiful; Munmun, Sarah; Mohiduzzaman, Md; Longvah, Thingnganing

    2016-12-15

    Concentrations of standard amino acids were determined in the composite samples (representing 30 agro-ecological zones of Bangladesh) of six prioritized key dietary protein sources: Oryza sativa (rice), Triticum aestivum (wheat flour), Lens culinaris (lentils), Pangusius pangusius (pangas), Labeo rohita (rohu) and Oreochromis mossambicus (tilapia). Digestible indispensable amino acid scores (DIAAS) was calculated using published data on amino acids' digestibility to evaluate the protein quality of these foods. Indispensable amino acid (IAA) contents (mg IAA/g protein), found to be highest in pangas (430) and lowest in wheat (336), of all these analyzed foods exceeded the FAO recommended daily allowance (277mg IAA/g protein) and contributed on average 40% to total amino acid contents. Untruncated DIAAS values ranged from 51% (lysine) in wheat to 106% (histidine) in pangas and distinguished pangas, rohu, and tilapia containing 'excellent quality' protein (DIAAS>100%) with potential to complement lower quality protein of cereals, fruits, and vegetables. PMID:27451158

  3. Association between serum folic acid level and erectile dysfunction.

    PubMed

    Karabakan, M; Erkmen, A E; Guzel, O; Aktas, B K; Bozkurt, A; Akdemir, S

    2016-06-01

    This study measured the serum folic acid (FA) level in patients with erectile dysfunction (ED) and evaluated the possible association between the serum FA level and erectile function. The study divided 120 patients with ED into 3 groups of 40 patients each: those with severe, moderate and mild ED. Forty healthy men served as controls. Fasting serum samples were obtained, and the total testosterone, cholesterol and FA levels were measured using chemiluminescent immunoassays. There were no significant differences in the mean age, mean body mass index or mean serum total testosterone and cholesterol levels among the three ED groups and controls (P > 0.05). The mean serum FA concentrations were 7.2 ± 3.7, 7.1 ± 3.2, 10.2 ± 4.6 and 10.7 ± 4.6 ng ml(-1) in the severe, moderate and mild ED and control groups respectively. The mean serum FA concentration was significantly higher in the control group than in the severe and moderate ED groups (both P < 0.001), but not the mild ED group (P = 0.95). Considering the significant differences in the serum FA levels between the control and ED groups, serum FA deficiency might reflect the severity of ED.

  4. Ocean acidification increases fatty acids levels of larval fish.

    PubMed

    Díaz-Gil, Carlos; Catalán, Ignacio A; Palmer, Miquel; Faulk, Cynthia K; Fuiman, Lee A

    2015-07-01

    Rising levels of anthropogenic carbon dioxide in the atmosphere are acidifying the oceans and producing diverse and important effects on marine ecosystems, including the production of fatty acids (FAs) by primary producers and their transfer through food webs. FAs, particularly essential FAs, are necessary for normal structure and function in animals and influence composition and trophic structure of marine food webs. To test the effect of ocean acidification (OA) on the FA composition of fish, we conducted a replicated experiment in which larvae of the marine fish red drum (Sciaenops ocellatus) were reared under a climate change scenario of elevated CO2 levels (2100 µatm) and under current control levels (400 µatm). We found significantly higher whole-body levels of FAs, including nine of the 11 essential FAs, and altered relative proportions of FAs in the larvae reared under higher levels of CO2. Consequences of this effect of OA could include alterations in performance and survival of fish larvae and transfer of FAs through food webs. PMID:26179801

  5. Ocean acidification increases fatty acids levels of larval fish

    PubMed Central

    Díaz-Gil, Carlos; Catalán, Ignacio A.; Palmer, Miquel; Faulk, Cynthia K.; Fuiman, Lee A.

    2015-01-01

    Rising levels of anthropogenic carbon dioxide in the atmosphere are acidifying the oceans and producing diverse and important effects on marine ecosystems, including the production of fatty acids (FAs) by primary producers and their transfer through food webs. FAs, particularly essential FAs, are necessary for normal structure and function in animals and influence composition and trophic structure of marine food webs. To test the effect of ocean acidification (OA) on the FA composition of fish, we conducted a replicated experiment in which larvae of the marine fish red drum (Sciaenops ocellatus) were reared under a climate change scenario of elevated CO2 levels (2100 µatm) and under current control levels (400 µatm). We found significantly higher whole-body levels of FAs, including nine of the 11 essential FAs, and altered relative proportions of FAs in the larvae reared under higher levels of CO2. Consequences of this effect of OA could include alterations in performance and survival of fish larvae and transfer of FAs through food webs. PMID:26179801

  6. Ocean acidification increases fatty acids levels of larval fish.

    PubMed

    Díaz-Gil, Carlos; Catalán, Ignacio A; Palmer, Miquel; Faulk, Cynthia K; Fuiman, Lee A

    2015-07-01

    Rising levels of anthropogenic carbon dioxide in the atmosphere are acidifying the oceans and producing diverse and important effects on marine ecosystems, including the production of fatty acids (FAs) by primary producers and their transfer through food webs. FAs, particularly essential FAs, are necessary for normal structure and function in animals and influence composition and trophic structure of marine food webs. To test the effect of ocean acidification (OA) on the FA composition of fish, we conducted a replicated experiment in which larvae of the marine fish red drum (Sciaenops ocellatus) were reared under a climate change scenario of elevated CO2 levels (2100 µatm) and under current control levels (400 µatm). We found significantly higher whole-body levels of FAs, including nine of the 11 essential FAs, and altered relative proportions of FAs in the larvae reared under higher levels of CO2. Consequences of this effect of OA could include alterations in performance and survival of fish larvae and transfer of FAs through food webs.

  7. Effects of protein deficiency and food restriction on lung ascorbic acid and glutathione in rats exposed to ozone

    SciTech Connect

    Dubick, M.A.; Heng, H.; Rucker, R.B.

    1985-08-01

    Weanling (52 +/- 4 g) or adult (259 +/- 16 g) male Sprague-Dawley rats were fed ad libitum casein-based diets containing 4 or 16% protein. A third group (food restricted) was fed daily the 16% protein diet, but at the food intake level of the 4% protein group. After 3 wk (weanling) or 5 wk (adults), half of the rats in each group were continuously exposed to 0.64 ppm ozone for 7 d. Ascorbic acid and reduced glutathione levels were then measured. In the heart and liver from weanling rats, ascorbic acid concentrations were lower in the protein-deficient group than in either control group. In the liver from weanling rats glutathione concentrations were also reduced in response to protein deficiency. Exposure to ozone produced no additional response. For adult rats the response for liver glutathione was similar to that of the weanlings. The liver ascorbate concentration, however, was consistently lower in adult rats compared to weanlings exposed to ozone. In lungs from adult rats, the ascorbic acid concentration was lower in the protein-deficient group than in either control group. On a whole-organ basis, both ascorbic acid and glutathione were usually higher in lungs from rats exposed to ozone than from those exposed to air. Interestingly, protein deficiency did not appear to compromise the lung's ability to maintain, in relative terms, the ascorbic acid or glutathione concentration in response to ozone.

  8. UCP2, a mitochondrial protein regulated at multiple levels.

    PubMed

    Donadelli, Massimo; Dando, Ilaria; Fiorini, Claudia; Palmieri, Marta

    2014-04-01

    An ever-increasing number of studies highlight the role of uncoupling protein 2 (UCP2) in a broad range of physiological and pathological processes. The knowledge of the molecular mechanisms of UCP2 regulation is becoming fundamental in both the comprehension of UCP2-related physiological events and the identification of novel therapeutic strategies based on UCP2 modulation. The study of UCP2 regulation is a fast-moving field. Recently, several research groups have made a great effort to thoroughly understand the various molecular mechanisms at the basis of UCP2 regulation. In this review, we describe novel findings concerning events that can occur in a concerted manner at various levels: Ucp2 gene mutation (single nucleotide polymorphisms), UCP2 mRNA and protein expression (transcriptional, translational, and protein turn-over regulation), UCP2 proton conductance (ligands and post-transcriptional modifications), and nutritional and pharmacological regulation of UCP2.

  9. Low copper and high manganese levels in prion protein plaques

    USGS Publications Warehouse

    Johnson, Christopher J.; Gilbert, P.U.P.A.; Abrecth, Mike; Baldwin, Katherine L.; Russell, Robin E.; Pedersen, Joel A.; McKenzie, Debbie

    2013-01-01

    Accumulation of aggregates rich in an abnormally folded form of the prion protein characterize the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs). The molecular triggers of plaque formation and neurodegeneration remain unknown, but analyses of TSE-infected brain homogenates and preparations enriched for abnormal prion protein suggest that reduced levels of copper and increased levels of manganese are associated with disease. The objectives of this study were to: (1) assess copper and manganese levels in healthy and TSE-infected Syrian hamster brain homogenates; (2) determine if the distribution of these metals can be mapped in TSE-infected brain tissue using X-ray photoelectron emission microscopy (X-PEEM) with synchrotron radiation; and (3) use X-PEEM to assess the relative amounts of copper and manganese in prion plaques in situ. In agreement with studies of other TSEs and species, we found reduced brain levels of copper and increased levels of manganese associated with disease in our hamster model. We also found that the in situ levels of these metals in brainstem were sufficient to image by X-PEEM. Using immunolabeled prion plaques in directly adjacent tissue sections to identify regions to image by X-PEEM, we found a statistically significant relationship of copper-manganese dysregulation in prion plaques: copper was depleted whereas manganese was enriched. These data provide evidence for prion plaques altering local transition metal distribution in the TSE-infected central nervous system.

  10. Low Copper and High Manganese Levels in Prion Protein Plaques

    PubMed Central

    Johnson, Christopher J.; Gilbert, P.U.P.A.; Abrecht, Mike; Baldwin, Katherine L.; Russell, Robin E.; Pedersen, Joel A.; Aiken, Judd M.; McKenzie, Debbie

    2013-01-01

    Accumulation of aggregates rich in an abnormally folded form of the prion protein characterize the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs). The molecular triggers of plaque formation and neurodegeneration remain unknown, but analyses of TSE-infected brain homogenates and preparations enriched for abnormal prion protein suggest that reduced levels of copper and increased levels of manganese are associated with disease. The objectives of this study were to: (1) assess copper and manganese levels in healthy and TSE-infected Syrian hamster brain homogenates; (2) determine if the distribution of these metals can be mapped in TSE-infected brain tissue using X-ray photoelectron emission microscopy (X-PEEM) with synchrotron radiation; and (3) use X-PEEM to assess the relative amounts of copper and manganese in prion plaques in situ. In agreement with studies of other TSEs and species, we found reduced brain levels of copper and increased levels of manganese associated with disease in our hamster model. We also found that the in situ levels of these metals in brainstem were sufficient to image by X-PEEM. Using immunolabeled prion plaques in directly adjacent tissue sections to identify regions to image by X-PEEM, we found a statistically significant relationship of copper-manganese dysregulation in prion plaques: copper was depleted whereas manganese was enriched. These data provide evidence for prion plaques altering local transition metal distribution in the TSE-infected central nervous system. PMID:23435237

  11. Low copper and high manganese levels in prion protein plaques.

    PubMed

    Johnson, Christopher J; Gilbert, P U P A; Abrecht, Mike; Baldwin, Katherine L; Russell, Robin E; Pedersen, Joel A; Aiken, Judd M; McKenzie, Debbie

    2013-02-01

    Accumulation of aggregates rich in an abnormally folded form of the prion protein characterize the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs). The molecular triggers of plaque formation and neurodegeneration remain unknown, but analyses of TSE-infected brain homogenates and preparations enriched for abnormal prion protein suggest that reduced levels of copper and increased levels of manganese are associated with disease. The objectives of this study were to: (1) assess copper and manganese levels in healthy and TSE-infected Syrian hamster brain homogenates; (2) determine if the distribution of these metals can be mapped in TSE-infected brain tissue using X-ray photoelectron emission microscopy (X-PEEM) with synchrotron radiation; and (3) use X-PEEM to assess the relative amounts of copper and manganese in prion plaques in situ. In agreement with studies of other TSEs and species, we found reduced brain levels of copper and increased levels of manganese associated with disease in our hamster model. We also found that the in situ levels of these metals in brainstem were sufficient to image by X-PEEM. Using immunolabeled prion plaques in directly adjacent tissue sections to identify regions to image by X-PEEM, we found a statistically significant relationship of copper-manganese dysregulation in prion plaques: copper was depleted whereas manganese was enriched. These data provide evidence for prion plaques altering local transition metal distribution in the TSE-infected central nervous system.

  12. Life-history evolution at the molecular level: adaptive amino acid composition of avian vitellogenins

    PubMed Central

    Hughes, Austin L.

    2015-01-01

    Avian genomes typically encode three distinct vitellogenin (VTG) egg yolk proteins (VTG1, VTG2 and VTG3), which arose by gene duplication prior to the most recent common ancestor of birds. Analysis of VTG sequences from 34 avian species in a phylogenetic framework supported the hypothesis that VTG amino acid composition has co-evolved with embryo incubation time. Embryo incubation time was positively correlated with the proportions of dietary essential amino acids (EAAs) in VTG1 and VTG2, and with the proportion of sulfur-containing amino acids in VTG3. These patterns were seen even when only semi-altricial and/or altricial species were considered, suggesting that the duration of embryo incubation is a major selective factor on the amino acid composition of VTGs, rather than developmental mode alone. The results are consistent with the hypothesis that the level of EAAs provided to the egg represents an adaptation to the loss of amino acids through breakdown over the course of incubation and imply that life-history phenotypes and VTG amino acid composition have co-evolved throughout the evolutionary history of birds. PMID:26224713

  13. Expression Pattern of Fatty Acid Binding Proteins in Celiac Disease Enteropathy

    PubMed Central

    Bottasso Arias, Natalia M.; García, Marina; Bondar, Constanza; Guzman, Luciana; Redondo, Agustina; Chopita, Nestor; Córsico, Betina; Chirdo, Fernando G.

    2015-01-01

    Celiac disease (CD) is an immune-mediated enteropathy that develops in genetically susceptible individuals following exposure to dietary gluten. Severe changes at the intestinal mucosa observed in untreated CD patients are linked to changes in the level and in the pattern of expression of different genes. Fully differentiated epithelial cells express two isoforms of fatty acid binding proteins (FABPs): intestinal and liver, IFABP and LFABP, respectively. These proteins bind and transport long chain fatty acids and also have other important biological roles in signaling pathways, particularly those related to PPARγ and inflammatory processes. Herein, we analyze the serum levels of IFABP and characterize the expression of both FABPs at protein and mRNA level in small intestinal mucosa in severe enteropathy and normal tissue. As a result, we observed higher levels of circulating IFABP in untreated CD patients compared with controls and patients on gluten-free diet. In duodenal mucosa a differential FABPs expression pattern was observed with a reduction in mRNA levels compared to controls explained by the epithelium loss in severe enteropathy. In conclusion, we report changes in FABPs' expression pattern in severe enteropathy. Consequently, there might be alterations in lipid metabolism and the inflammatory process in the small intestinal mucosa. PMID:26346822

  14. Oleic acid enhances G protein coupled receptor 43 expression in bovine intramuscular adipocytes but not in subcutaneous adipocytes.

    PubMed

    Chung, K Y; Smith, S B; Choi, S H; Johnson, B J

    2016-05-01

    We hypothesized that fatty acids would differentially affect G protein coupled receptor (GPR) 43 mRNA expression and GPR43 protein concentrations in bovine intramuscular (IM) and subcutaneous (SC) adipocytes. The GPR43 protein was detected in bovine liver, pancreas, and semimembranosus (MUS) muscle in samples taken at slaughter. Similarly, GPR43 protein levels were similar in IM adipose tissue and SM muscle but was barely detectable in SC adipose tissue. Primary cultures of IM and SC stromal vascular cells were isolated from bovine adipose tissues. Oleic acid (100 μ) stimulated PPARγ gene expression and decreased stearoyl-CoA desaturase (SCD) gene expression but had no effect on GPR43 gene expression, which was readily detectable in both IM and SC adipocytes. Differentiation cocktail (Diff; 10 μ insulin, 4 μ dexamethasone, and 10 μ ciglitizone) stimulated CCAAT/enhancer-binding protein β (C/EBPβ) and PPARγ gene expression in SC but not IM adipocytes, but Diff increased SCD gene expression in both cell types. Linoleic acid (10 µ) increased PPARγ gene expression relative to Diff cocktail in SC adipocytes, whereas linoleic acid and α-linolenic decreased SCD gene expression relative to control adipocytes and adipocytes incubated with Diff ( < 0.05). Increasing concentrations of oleic acid (1, 10, 100, and 500 μM) increased GPR43 protein and mRNA expression in IM but not SC adipocytes. These data indicated that oleic acid alters mRNA and protein concentrations of GPR43 in bovine IM adipocytes. PMID:27285685

  15. Identification of a family of fatty acid-speciated Sonic Hedgehog proteins, whose members display differential biological properties

    PubMed Central

    Houel, Stephane; Rodgriguez-Blanco, Jezabel; Singh, Samer; Schilling, Neal; J.Capobianco, Anthony; Ahn, Natalie G.; Robbins, David J.

    2015-01-01

    SUMMARY Hedgehog (HH) proteins are proteolytically processed into a biologically active form, which is covalently modified by cholesterol and palmitate. However, most studies of HH biogenesis have characterized protein from cells in which HH is over-expressed. We purified Sonic Hedgehog (SHH) from cells expressing physiologically relevant levels, and showed that it was more potent than SHH isolated from over-expressing cells. Furthermore, the SHH in our preparations were modified with a diverse spectrum of fatty acids on their amino-termini, and this spectrum of fatty acids varied dramatically depending on the growth conditions of the cells. The fatty acid composition of SHH affected its trafficking to lipid rafts, as well as its potency. Our results suggest that HH proteins exist as a family of diverse lipid-speciated proteins, which might be altered in different physiological and pathological contexts to regulate distinct properties of HH proteins. PMID:25732819

  16. Photolabeling of brain membrane proteins by lysergic acid diethylamide

    SciTech Connect

    Mahon, A.C.; Hartig, P.R.

    1982-04-05

    /sup 3/H-Lysergic acid diethylamide (/sup 3/H-LSD) is irreversibly incorporated into bovine caudate membranes during ultraviolet light illumination. The incorporated radioligand apparently forms a covalent bond with a sub-population of the membrane proteins. Although the photolabeling pattern differs significantly from the Coomassie blue staining pattern on SDS gels, the photolabeling is apparently not specific for LSD binding sites associated with neurotransmitter receptors. /sup 3/H-LSD photolabeling can occur during prolonged exposure of membrane samples to room lighting and thus may introduce artifacts into receptor binding assays.

  17. Acid-base chemistry of frustrated water at protein interfaces.

    PubMed

    Fernández, Ariel

    2016-01-01

    Water molecules at a protein interface are often frustrated in hydrogen-bonding opportunities due to subnanoscale confinement. As shown, this condition makes them behave as a general base that may titrate side-chain ammonium and guanidinium cations. Frustration-based chemistry is captured by a quantum mechanical treatment of proton transference and shown to remove same-charge uncompensated anticontacts at the interface found in the crystallographic record and in other spectroscopic information on the aqueous interface. Such observations are untenable within classical arguments, as hydronium is a stronger acid than ammonium or guanidinium. Frustration enables a directed Grotthuss mechanism for proton transference stabilizing same-charge anticontacts.

  18. Structure and function analysis of protein-nucleic acid complexes

    NASA Astrophysics Data System (ADS)

    Kuznetsova, S. A.; Oretskaya, T. S.

    2016-05-01

    The review summarizes published data on the results and achievements in the field of structure and function analysis of protein-nucleic acid complexes by means of main physical and biochemical methods, including X-ray diffraction, nuclear magnetic resonance spectroscopy, electron and atomic force microscopy, small-angle X-ray and neutron scattering, footprinting and cross-linking. Special attention is given to combined approaches. The advantages and limitations of each method are considered, and the prospects of their application for wide-scale structural studies in vivo are discussed. The bibliography includes 145 references.

  19. Acid-base chemistry of frustrated water at protein interfaces.

    PubMed

    Fernández, Ariel

    2016-01-01

    Water molecules at a protein interface are often frustrated in hydrogen-bonding opportunities due to subnanoscale confinement. As shown, this condition makes them behave as a general base that may titrate side-chain ammonium and guanidinium cations. Frustration-based chemistry is captured by a quantum mechanical treatment of proton transference and shown to remove same-charge uncompensated anticontacts at the interface found in the crystallographic record and in other spectroscopic information on the aqueous interface. Such observations are untenable within classical arguments, as hydronium is a stronger acid than ammonium or guanidinium. Frustration enables a directed Grotthuss mechanism for proton transference stabilizing same-charge anticontacts. PMID:26762189

  20. What is the relationship between gestational age and docosahexaenoic acid (DHA) and arachidonic acid (ARA) levels?

    PubMed

    Baack, Michelle L; Puumala, Susan E; Messier, Stephen E; Pritchett, Deborah K; Harris, William S

    2015-09-01

    Long chain polyunsaturated fatty acids (LCPUFA) including docosahexaenoic acid (DHA) and arachidonic acid (ARA) are increasingly transferred from mother to fetus late in pregnancy. Infants born before this transfer is complete are at risk for deficiency. This study determines the relationship between gestational age (GA) and circulating LCPUFA levels to better understand the unique needs of premature infants born at various GAs. Whole blood was collected within the first 7 days of life from 60 preterm (≤34 weeks GA) and 30 term infants (≥38 weeks GA) and FA levels were analyzed. Since concurrent intravenous lipid emulsion can skew composition data, blood LCPUFA concentrations were also measured. Levels were compared among groups, and linear regression models were used to examine the association between FA composition and GA. Preterm infants had significantly lower DHA and ARA levels than term peers, and whether assessed as concentrations or compositions, both directly correlated with GA (p<0.0001). Moreover, FA comparisons suggest that premature infants have impaired synthesis of LCPUFAs from precursors and may require preformed DHA and ARA. This study confirms that essential FA status is strongly related to GA, and that those babies born the earliest are at the greatest risk of LCPUFA deficiency.

  1. What is the Relationship between Gestational Age and Docosahexaenoic Acid (DHA) and Arachidonic Acid (ARA) Levels?

    PubMed Central

    Baack, Michelle L; Puumala, Susan E; Messier, Stephen E; Pritchett, Deborah K; Harris, William S

    2015-01-01

    Long chain polyunsaturated fatty acids (LCPUFA) including docosahexaenoic acid (DHA) and arachidonic acid (ARA) are increasingly transferred from mother to fetus late in pregnancy. Infants born before this transfer is complete are at risk for deficiency. This study determines the relationship between gestational age (GA) and circulating LCPUFA levels to better understand the unique needs of premature infants born at various GAs. Whole blood was collected within the first 7 days of life from 60 preterm (≤34 weeks GA) and 30 term infants (≥38 weeks GA) and FA levels were analyzed. Since concurrent intravenous lipid emulsion can skew composition data, blood LCPUFA concentrations were also measured. Levels were compared among groups, and linear regression models were used to examine the association between FA composition and GA. Preterm infants had significantly lower DHA and ARA levels than term peers, and whether assessed as concentrations or compositions, both directly correlated with GA (p<0.0001). Moreover, FA comparisons suggest that premature infants have impaired synthesis of LCPUFAs from precursors and may require preformed DHA and ARA. This study confirms that essential FA status is strongly related to GA, and that those babies born the earliest are at the greatest risk of LCPUFA deficiency. PMID:26205427

  2. Role of protein and ferulic acid in the emulsification properties of sugar beet pectin.

    PubMed

    Siew, Chee Kiong; Williams, Peter A

    2008-06-11

    The ability of sugar beet pectin to stabilize 20% w/w limonene oil-in-water emulsions has been investigated. The size of the oil droplets as determined by laser diffraction measurements decreased from about 15 mum to about 6 mum when the pectin concentration increased from 0.05% to 2% w/w but leveled off thereafter, suggesting complete coverage of the oil droplets by the polymer at this optimum concentration. Isotherms for the adsorption of pectin, protein, and ferulic acid were constructed. The adsorption capacities at the oil-water interface of approximately 1.4 and approximately 0.2 mg/m (2) for protein and ferulic acid, respectively, compared to approximately 9.5 mg/m (2) for pectin revealed that the adsorbed fractions of the pectin sample were rich in protein (14.7%) and ferulic acid (2.1%) given that there were only 2.7% protein and 1.06% ferulic acid present in the whole pectin sample. Direct measurements on the adsorbed fraction recovered from the oil droplets via desorption with SDS confirmed that it contained 11.1% protein and 2.16% ferulic acid. The results suggest that one or both of these two functional groups adsorb onto the surface of the oil droplets and stabilize the emulsions. High molecular mass fractions adsorbed preferentially onto oil droplets during emulsification. As compared to those made with gum arabic, the emulsion samples made with sugar beet pectin samples exhibited similar (or even slightly higher) stability.

  3. Separation of membranes from acid-solubilized fish muscle proteins with the aid of calcium ions and organic acids.

    PubMed

    Liang, Yong; Hultin, Herbert O

    2005-04-20

    Calcium chloride, and to a lesser extent MgCl2, aided in the separation of membranes by centrifugation from cod (Gadus morhua) muscle homogenates solubilized at pH 3 in the presence of citric acid or malic acid but not lactic acid. Adding citric acid and Ca2+ before solubilizing the cod muscle homogenates was needed for the effect. At 1 mM citric acid, 70-80% of the phospholipid and 25-30% of the protein were removed at 10 mM Ca2+. At 8 mM Ca2+, citric acid showed an optimal effect on phospholipid removal at 5 mM with 90% of the phospholipid and 35% of the protein removed. The treatment with citric acid and Ca2+ was also effective in separating the membrane from solubilized herring (Clupea harengus) muscle homogenate. Ca2+ and citric acid might exert their influence by disconnecting linkages between membranes and cytoskeletal proteins.

  4. Residue level quantification of protein stability in living cells.

    PubMed

    Monteith, William B; Pielak, Gary J

    2014-08-01

    The intracellular milieu differs from the dilute conditions in which most biophysical and biochemical studies are performed. This difference has led both experimentalists and theoreticians to tackle the challenging task of understanding how the intracellular environment affects the properties of biopolymers. Despite a growing number of in-cell studies, there is a lack of quantitative, residue-level information about equilibrium thermodynamic protein stability under nonperturbing conditions. We report the use of NMR-detected hydrogen-deuterium exchange of quenched cell lysates to measure individual opening free energies of the 56-aa B1 domain of protein G (GB1) in living Escherichia coli cells without adding destabilizing cosolutes or heat. Comparisons to dilute solution data (pH 7.6 and 37 °C) show that opening free energies increase by as much as 1.14 ± 0.05 kcal/mol in cells. Importantly, we also show that homogeneous protein crowders destabilize GB1, highlighting the challenge of recreating the cellular interior. We discuss our findings in terms of hard-core excluded volume effects, charge-charge GB1-crowder interactions, and other factors. The quenched lysate method identifies the residues most important for folding GB1 in cells, and should prove useful for quantifying the stability of other globular proteins in cells to gain a more complete understanding of the effects of the intracellular environment on protein chemistry.

  5. Molecular evolution of monotreme and marsupial whey acidic protein genes.

    PubMed

    Sharp, Julie A; Lefèvre, Christophe; Nicholas, Kevin R

    2007-01-01

    Whey acidic protein (WAP), a major whey protein present in milk of a number of mammalian species has characteristic cysteine-rich domains known as four-disulfide cores (4-DSC). Eutherian WAP, expressed in the mammary gland throughout lactation, has two 4-DSC domains, (DI-DII) whereas marsupial WAP, expressed only during mid-late lactation, contains an additional 4-DSC (DIII), and has a DIII-D1-DII configuration. We report the expression and evolution of echidna (Tachyglossus aculeatus) and platypus (Onithorhynchus anatinus) WAP cDNAs. Predicted translation of monotreme cDNAs showed echidna WAP contains two 4-DSC domains corresponding to DIII-DII, whereas platypus WAP contains an additional domain at the C-terminus with homology to DII and has the configuration DIII-DII-DII. Both monotreme WAPs represent new WAP protein configurations. We propose models for evolution of the WAP gene in the mammalian lineage either through exon loss from an ancient ancestor or by rapid evolution via the process of exon shuffling. This evolutionary outcome may reflect differences in lactation strategy between marsupials, monotremes, and eutherians, and give insight to biological function of the gene products. WAP four-disulfide core domain 2 (WFDC2) proteins were also identified in echidna, platypus and tammar wallaby (Macropus eugenii) lactating mammary cells. WFDC2 proteins are secreted proteins not previously associated with lactation. Mammary gland expression of tammar WFDC2 during the course of lactation showed WFDC2 was elevated during pregnancy, reduced in early lactation and absent in mid-late lactation.

  6. Bioprocess monitoring: minimizing sample matrix effects for total protein quantification with bicinchoninic acid assay.

    PubMed

    Reichelt, Wieland N; Waldschitz, Daniel; Herwig, Christoph; Neutsch, Lukas

    2016-09-01

    Determining total protein content is a routine operation in many laboratories. Despite substantial work on assay optimization interferences, the widely used bicinchoninic acid (BCA) assay remains widely recognized for its robustness. Especially in the field of bioprocess engineering the inaccuracy caused by interfering substances remains hardly predictable and not well understood. Since the introduction of the assay, sample pre-treatment by trichloroacetic acid (TCA) precipitation has been indicated as necessary and sufficient to minimize interferences. However, the sample matrix in cultivation media is not only highly complex but also dynamically changing over process time in terms of qualitative and quantitative composition. A significant misestimation of the total protein concentration of bioprocess samples is often observed when following standard work-up schemes such as TCA precipitation, indicating that this step alone is not an adequate means to avoid measurement bias. Here, we propose a modification of the BCA assay, which is less influenced by sample complexity. The dynamically changing sample matrix composition of bioprocessing samples impairs the conventional approach of compensating for interfering substances via a static offset. Hence, we evaluated the use of a correction factor based on an internal spike measurement for the respective samples. Using protein spikes, the accuracy of the BCA protein quantification could be improved fivefold, taking the BCA protein quantification to a level of accuracy comparable to other, more expensive methods. This will allow reducing expensive iterations in bioprocess development to due inaccurate total protein analytics. PMID:27314233

  7. Dietary aspartame with protein on plasma and brain amino acids, brain monoamines and behavior in rats.

    PubMed

    Torii, K; Mimura, T; Takasaki, Y; Ichimura, M

    1986-01-01

    Aspartame (APM; L-aspartyl-L-phenylalanine methyl ester), was investigated for its ability to alter levels of the large neutral amino acids and monoamines in overnight fasted rats allowed to consume meals with or without protein for two hours. Additionally, the possible long term behavioral consequences of APM in 25% casein diets with or without 10% sucrose were determined. Acute APM ingestion increased both plasma and brain phenylalanine and tyrosine levels, but brain tryptophan levels were not altered regardless of dietary protein. Brain norepinephrine and dopamine levels were unaltered by any of the diet while serotonin levels were slightly increased when a protein-free diet was consumed. But APM and/or protein ingestion minimized this increase of brain serotonin levels as much as controls. Chronic APM ingestion failed to influence diurnal feeding patterns, meal size distributions, or diurnal patterns of spontaneous motor activity. The chronic ingestion of abuse doses of APM produced no significant chemical changes in brain capable of altering behavioral parameters believed to be controlled by monoamines in rats.

  8. Serum and Urinary Malondialdehyde (MDA), Uric acid, and Protein as markers of perinatal asphyxia

    PubMed Central

    El Bana, Sawsan Mahmoud; Maher, Sheren Esam; Gaber, Amani Fawzy; Aly, Sanaa Shaker

    2016-01-01

    Introduction Perinatal asphyxia (PA) is among the leading causes of neonatal morbidity and death in neonatal intensive care units (NICUs). The aims of this research were to determine the concentrations of malondialdehyde (MDA), urine MDA, uric acid, and protein in the cord blood of neonates with perinatal asphyxia and to determine their relationship with the severity of perinatal asphyxia. Methods This matched case-control study was conducted from October 2012 to March 2013. All of the cases and controls were selected from the Gynecology & Obstetrics Department and the NICUs, at Qous Central Hospital in Qena, Egypt. We allocated 20 full-term neonates who had perinatal asphyxia to the case group. Also, we selected 20 healthy neonates for the control group. The subjects were matched with respect to age and gender. At birth and 48 hours later, measurements were made of MDA in cord blood and urine, and uric acid, protein, and creatinine also were measured in both groups. The data were analyzed by SPSS, version 17, using the independent-samples t-test, ANOVA, Tukey’s test, and Spearman’s correlation coefficient. Results At birth and 48 hr later, the newborns’ with PA had significantly higher levels of MDA in the cord blood, mean urinary uric acid/creatinine (UUA:Cr), protein/creatinine (UP:Cr), and MDA/creatinine ratio (UMDA:Cr) than the controls; their PA levels were correlated with the degree of hypoxic-ischemic encephalopathy (HIE). The babies who died due to PA had significantly higher levels of cord blood MDA, and they also had higher UUA:Cr, UP:Cr, and UMDA:Cr ratios than the babies who survived. Conclusion The concentration of MDA in cord blood can be used as a diagnostic marker of oxidative stress in asphyxiated neonates. The ratios of the urinary excretion rates of uric acid, protein, and MDA to creatinine increased as the severity of perinatal asphyxia and associated brain damage increased. PMID:27648187

  9. Serum and Urinary Malondialdehyde (MDA), Uric acid, and Protein as markers of perinatal asphyxia

    PubMed Central

    El Bana, Sawsan Mahmoud; Maher, Sheren Esam; Gaber, Amani Fawzy; Aly, Sanaa Shaker

    2016-01-01

    Introduction Perinatal asphyxia (PA) is among the leading causes of neonatal morbidity and death in neonatal intensive care units (NICUs). The aims of this research were to determine the concentrations of malondialdehyde (MDA), urine MDA, uric acid, and protein in the cord blood of neonates with perinatal asphyxia and to determine their relationship with the severity of perinatal asphyxia. Methods This matched case-control study was conducted from October 2012 to March 2013. All of the cases and controls were selected from the Gynecology & Obstetrics Department and the NICUs, at Qous Central Hospital in Qena, Egypt. We allocated 20 full-term neonates who had perinatal asphyxia to the case group. Also, we selected 20 healthy neonates for the control group. The subjects were matched with respect to age and gender. At birth and 48 hours later, measurements were made of MDA in cord blood and urine, and uric acid, protein, and creatinine also were measured in both groups. The data were analyzed by SPSS, version 17, using the independent-samples t-test, ANOVA, Tukey’s test, and Spearman’s correlation coefficient. Results At birth and 48 hr later, the newborns’ with PA had significantly higher levels of MDA in the cord blood, mean urinary uric acid/creatinine (UUA:Cr), protein/creatinine (UP:Cr), and MDA/creatinine ratio (UMDA:Cr) than the controls; their PA levels were correlated with the degree of hypoxic-ischemic encephalopathy (HIE). The babies who died due to PA had significantly higher levels of cord blood MDA, and they also had higher UUA:Cr, UP:Cr, and UMDA:Cr ratios than the babies who survived. Conclusion The concentration of MDA in cord blood can be used as a diagnostic marker of oxidative stress in asphyxiated neonates. The ratios of the urinary excretion rates of uric acid, protein, and MDA to creatinine increased as the severity of perinatal asphyxia and associated brain damage increased.

  10. Survival of Listeria innocua in rainbow trout protein recovered by isoelectric solubilization and precipitation with acetic and citric acids.

    PubMed

    Otto, R A; Paker, I; Bane, L; Beamer, S; Jaczynski, J; Matak, K E

    2011-08-01

    During mechanical fish processing, a substantial amount of protein is discarded as by-products. Isoelectric solubilization and precipitation (ISP) is a process that uses extreme pH shifts to solubilize and precipitate protein from by-products to recover previously discarded protein. Typically, strong acids are used for pH reduction, but these acids do not have a pasteurization effect (6 log reduction) on bacterial load; therefore, organic acids were used during ISP processing to test the impact on Listeria innocua concentrations. Headed and gutted rainbow trout (Oncorhynchus mykiss) were inoculated with L. innocua, homogenized, and brought to the target pH with granular citric acid (pH 2.0 and 2.5) or glacial acetic acid (pH 3.0 and 3.5). Proteins were solubilized for 10 min at 4°C, and insoluble components (e.g., skin and insoluble protein) were removed by centrifugation. The remaining solution was pH shifted to the protein isoelectric point (pH 5.5) with sodium hydroxide, and precipitated protein was separated from the water. Microbial cells for each component (proteins, insolubles, and water) were enumerated on modified Oxford agar (MOX) and tryptic soy agar with 6% yeast extract (TSAYE). The sums of the surviving cells from each component were compared with the initial inoculum levels. No significant differences were observed between results obtained from TSAYE and from MOX (P > 0.05). Significant reductions in microbial populations were detected, regardless of pH or acid type (P < 0.05). The greatest reduction was at pH 3.0 with glacial acetic acid, resulting in a mean reduction of 6.41 log CFU/g in the recovered protein and 5.88 log CFU/g in the combined components. These results demonstrate the antimicrobial potential of organic acids in ISP processing.

  11. Changes in intramuscular amino acid levels in submaximally exercised horses - a pilot study.

    PubMed

    van den Hoven, R; Bauer, A; Hackl, S; Zickl, M; Spona, J; Zentek, J

    2010-08-01

    The time-dependent changes in intramuscular amino acid (AA) levels caused by exercise and by feeding a protein/AA supplement were analysed in nine horses. Horses were submitted to a total of four standardized exercise tests (SETs). Amino acid concentrations were determined prior to, immediately after, 4 and 18 h after exercise. The experiment was subdivided into two consecutive periods of 3 weeks. In each period two SETs were performed. In the second period, horses were given a protein/AA supplement within 1 h after exercise. Significant changes in mean plasma AA levels similar to previous studies were noted to be time-dependent and to be associated with feeding the supplement. The intramuscular concentrations of the free AA in relation to pre-exercise levels showed significant time-dependent changes for alanine, asparagine, aspartate, citrulline, glutamine, glycine, isoleucine, leucine, methionine, serine, taurine, threonine, tyrosine and valine. Feeding the supplement significantly increased the 4 h post-exercise intramuscular concentration of alanine, isoleucine, methionine and tyrosine. At 18 h after exercise, apart from isoleucine and methionine, levels were still increased and also those of asparagine, histidine and valine in relation to none treatment. Hence, it was concluded that AA mixtures administered orally to horses within 1 h after exercise increased intramuscular AA pool. PMID:19663973

  12. Ribonucleic Acid Regulation in Permeabilized Cells of Escherichia coli Capable of Ribonucleic Acid and Protein Synthesis1

    PubMed Central

    Atherly, Alan G.

    1974-01-01

    A cell permeabilization procedure is described that reduces viability less than 10% and does not significantly reduce the rates of ribonucleic acid and protein synthesis when appropriately supplemented. Permeabilization abolishes the normal stringent coupling of protein and ribonucleic acid synthesis. PMID:4364330

  13. The absence of protein--sparing effects utilizing crystalline amino acids in stressed patients.

    PubMed

    Ching, N; Mills, C J; Grossi, C; Angers, J W; Jham, G; Zurawinsky, H; Nealon, T F

    1979-11-01

    The protein-sparing effects of the peripheral infusion of crystalline amino acids (PAA) was studied metabolically in selected surgical patients subjected to various degrees of stress. Twenty-one patients (sixteen cancer patients receiving chemotherapy and/or radiotherapy, three with major abdominal traumatic injuries and four with paralytic ileus) were infused with 2 1/24 hours of a solution of 4.2% Travasol amino acids with only 5% glucose as a source of nonprotein calories. One-half of the cancer patients were also allowed ad libitum oral intake of a regular hospital diet or Vivonex-HN. The nutritional status was evaluated by measuring changes in body weight, serum albumin levels and nitrogen balance. Body weight decreased in only the trauma patients. When these solutions were the sole source of nutrients all patients were in negative nitrogen balance and had significant decreases in their serum albumin levels. Serum albumin levels were preserved only when extra sources of calories were provided. The infusion of the crystalline amino acids without adequate levels of nonprotein energy did not conserve protein in these stressed patients.

  14. Shedding light on proteins, nucleic acids, cells, humans and fish

    NASA Technical Reports Server (NTRS)

    Setlow, Richard B.

    2002-01-01

    I was trained as a physicist in graduate school. Hence, when I decided to go into the field of biophysics, it was natural that I concentrated on the effects of light on relatively simple biological systems, such as proteins. The wavelengths absorbed by the amino acid subunits of proteins are in the ultraviolet (UV). The wavelengths that affect the biological activities, the action spectra, also are in the UV, but are not necessarily parallel to the absorption spectra. Understanding these differences led me to investigate the action spectra for affecting nucleic acids, and the effects of UV on viruses and cells. The latter studies led me to the discovery of the important molecular nature of the damages affecting DNA (cyclobutane pyrimidine dimers) and to the discovery of nucleotide excision repair. Individuals with the genetic disease xeroderma pigmentosum (XP) are extraordinarily sensitive to sunlight-induced skin cancer. The finding, by James Cleaver, that their skin cells were defective in DNA repair strongly suggested that DNA damage was a key step in carcinogenesis. Such information was important for estimating the wavelengths in sunlight responsible for human skin cancer and for predicting the effects of ozone depletion on the incidence of non-melanoma skin cancer. It took experiments with backcross hybrid fish to call attention to the probable role of the longer UV wavelengths not absorbed by DNA in the induction of melanoma. These reflections trace the biophysicist's path from molecules to melanoma.

  15. Shedding light on proteins, nucleic acids, cells, humans and fish.

    PubMed

    Setlow, Richard B

    2002-03-01

    I was trained as a physicist in graduate school. Hence, when I decided to go into the field of biophysics, it was natural that I concentrated on the effects of light on relatively simple biological systems, such as proteins. The wavelengths absorbed by the amino acid subunits of proteins are in the ultraviolet (UV). The wavelengths that affect the biological activities, the action spectra, also are in the UV, but are not necessarily parallel to the absorption spectra. Understanding these differences led me to investigate the action spectra for affecting nucleic acids, and the effects of UV on viruses and cells. The latter studies led me to the discovery of the important molecular nature of the damages affecting DNA (cyclobutane pyrimidine dimers) and to the discovery of nucleotide excision repair. Individuals with the genetic disease xeroderma pigmentosum (XP) are extraordinarily sensitive to sunlight-induced skin cancer. The finding, by James Cleaver, that their skin cells were defective in DNA repair strongly suggested that DNA damage was a key step in carcinogenesis. Such information was important for estimating the wavelengths in sunlight responsible for human skin cancer and for predicting the effects of ozone depletion on the incidence of non-melanoma skin cancer. It took experiments with backcross hybrid fish to call attention to the probable role of the longer UV wavelengths not absorbed by DNA in the induction of melanoma. These reflections trace the biophysicist's path from molecules to melanoma.

  16. Protein utilization and amino acid digestibility of canola meal in response to phytase in broiler chickens.

    PubMed

    Kong, C; Adeola, O

    2011-07-01

    The regression method was used in a 14-d broiler chicken study to determine the true ileal digestibility of amino acid (AA) and protein utilization in canola meal (CM, 388 g of CP/kg) without or with added phytase. Experimental treatments consisted of 2 factors, phytase at 2 levels (0 or 1,500 phytase units/kg) and CM at 3 levels (125, 250, or 375 g/kg). Birds received a standard starter diet from d 1 to 8 and the assay diets from d 8 to 22 posthatch. On d 8, a total of 384 birds were allocated to 6 dietary treatments in a randomized complete block design; excreta were collected from d 12 to 14 and d 19 to 21, and ileal digesta were collected on d 22 posthatch. True ileal indispensable AA digestibility of CM were derived from the regression of AA flow (mg/kg of DM intake) at the terminal ileum against the intake of AA (mg/kg of dietary DM) of birds fed diets without or with phytase. Body weight gain (BWG), protein gain, and protein intake increased linearly (P < 0.001) with increasing CM level, regardless of phytase supplementation. Effects of phytase (P < 0.05) were observed on BWG and the protein efficiency ratio from d 8 to 15, whereas effects of phytase (P < 0.05) were observed on BWG and protein gain from d 15 to 22. There was no effect of phytase on protein intake and net protein utilization from d 8 to 22. Phytase supplementation at 1,500 phytase units/kg did not affect true ileal digestibility of any AA in CM. In conclusion, the results of this study indicated that phytase supplementation improved the protein efficiency ratio of birds fed diets containing CM as the sole protein source from d 8 to 15 posthatch but did not affect the true ileal digestibility of AA in CM as determined by the regression method.

  17. The intracellular Ca(2+)-pump inhibitors thapsigargin and cyclopiazonic acid induce stress proteins in mammalian chondrocytes.

    PubMed

    Cheng, T C; Benton, H P

    1994-07-15

    Primary cultures of mammalian articular chondrocytes respond to treatment with the intracellular Ca(2+)-pump inhibitors thapsigargin (TG) and cyclopiazonic acid by specific changes in protein synthesis consistent with a stress response. Two-dimensional gel electrophoresis of newly synthesized proteins confirmed that the response was consistent with the induction of glucose-regulated proteins. The effects of low-dose TG (10 nM), measured by changes in [35S]methionine labelling of newly synthesized proteins, can first be observed by 10 h and are maximal by 24 h. The pattern of changes induced by TG is shared with cyclopiazonic acid, but effects of both perturbants differ significantly from changes induced by heat shock. Upon removal of TG, normal protein synthesis is restored by 48 h. Immunoblots showed increased concentrations of the stress proteins HSP90, HSP72/73 and HSP60 in chondrocytes treated with TG, but induction of newly synthesized heat-shock proteins by TG was not apparent on [35S]methionine-labelled gels. The alterations in protein synthesis induced by Ca(2+)-pump inhibitors were unaffected by BAPTA-AM loading, which clamped cytosolic Ca2+ at resting levels. We conclude that inhibition of intracellular Ca(2+)-pump activity can elicit a stress response, which has important implications for the interpretation of chronic use of Ca(2+)-pump inhibitors. In particular, the activation of the cellular shock response should be considered in interpreting the regulation of protein synthesis and cell survival by Ca(2+)-pump inhibitors such as TG. PMID:8043004

  18. Accumulated p53 protein and UVA protection level of sunscreens.

    PubMed

    Seité, S; Moyal, D; Verdier, M P; Hourseau, C; Fourtanier, A

    2000-02-01

    Nuclear p53 expression is a sensitive parameter for the detection of ultraviolet (UV)-induced skin damage, and it has been used as an endpoint to evaluate the effectiveness of sunscreens. In this study, we compared the protection provided by two sunscreens having identical sun protection factors (SPF) but different UVA protection factors (UVA-PF) measured by the persistent pigment darkening method (PPD). The SPF of the sunscreens was 7 and the UVA-PF were respectively 7 and 3. Nuclear p53 protein was quantified in human skin biopsies treated with sunscreens and exposed 8 times to 5 MED of solar simulated radiation (SSR). The results showed that both sunscreens offered only partial protection against the increased expression of nuclear p53 protein induced by repetitive SSR exposures. However, a significantly lower level of p53-positive cells was found in areas protected with the sunscreen having the higher UVA-PF compared to the other sunscreen protected areas. In order to verify whether the difference in efficacy of these products was due to the difference in UVA absorption capacity, we quantified epidermal p53 protein accumulation after 8 exposures to either UVA (320-400 nm) or UVA1 (340-400 nm). We showed that as with SSR, repetitive exposures to 12.5 and 25 J/cm2 of UVA or UVA1 induced a significant increase in p53-positive cells in the human epidermis. These results confirmed that SPF determined on the basis of an acute erythemal reaction does not predict the level of protection against cumulative damage. They also showed that the protection provided by two sunscreens with different UVA protection factors is different (based on nuclear p53 protein accumulation), and that the PPD method can distinguish varying levels of sunscreen efficacy against UVA-induced cell damage. PMID:10721857

  19. Zebrafish cellular nucleic acid-binding protein: gene structure and developmental behaviour.

    PubMed

    Armas, Pablo; Cachero, Sebastián; Lombardo, Verónica A; Weiner, Andrea; Allende, Miguel L; Calcaterra, Nora B

    2004-08-01

    Here we analyse the structural organisation and expression of the zebrafish cellular nucleic acid-binding protein (zCNBP) gene and protein. The gene is organised in five exons and four introns. A noteworthy feature of the gene is the absence of a predicted promoter region. The coding region encodes a 163-amino acid polypeptide with the highly conserved general structural organisation of seven CCHC Zn knuckle domains and an RGG box between the first and the second Zn knuckles. Although theoretical alternative splicing is possible, only one form of zCNBP is actually detected. This form is able to bind to single-stranded DNA and RNA probes in vitro. The analysis of zCNBP developmental expression shows a high amount of CNBP-mRNA in ovary and during the first developmental stages. CNBP-mRNA levels decrease while early development progresses until the midblastula transition (MBT) stage and increases again thereafter. The protein is localised in the cytoplasm of blastomeres whereas it is mainly nuclear in developmental stages after the MBT. These findings suggest that CNBP is a strikingly conserved single-stranded nucleic acid-binding protein which might interact with maternal mRNA during its storage in the embryo cell cytoplasm. It becomes nuclear once MBT takes place possibly in order to modulate zygotic transcription and/or to associate with newly synthesised transcripts.

  20. Effect of protein level and protein source on zinc absorption in humans

    SciTech Connect

    Sandstroem, B.A.; Almgren, A.; Kivistoe, B.C.; Cederblad, A.

    1989-01-01

    The effect of increasing levels of various protein sources on zinc absorption from a legume-based meal was studied in humans with the use of a radionuclide technique. The meals were extrinsically labelled with 65Zn and absorption was determined from measurements of the whole-body retention of the isotope. The mean fractional zinc absorption for the 13 meals was 24.7 +/- 6.9% and was only influenced by the protein content of the meal to a limited extent (r = 0.45). However, the amount of zinc absorbed from the meals was strongly correlated with both the protein (r = 0.85) and zinc content (r = 0.86): 5.9 +/- 1.7 mumol of zinc was absorbed from the basal bean meal which had the lowest protein content; the addition of low zinc chicken doubled the protein content and increased zinc absorption to 10.3 +/- 2.0 mumol; the addition of zinc-rich beef also doubled the protein content, however, zinc absorption was increased to 15.9 +/- 4.7 mumol. It is concluded that the zinc content of the main protein source of the diet determines the amount of zinc absorbed to a large extent. However, relatively small amounts of animal protein can significantly improve the value of a legume-based meal as a source of zinc.

  1. Ascertaining effects of nanoscale polymeric interfaces on competitive protein adsorption at the individual protein level.

    PubMed

    Song, Sheng; Xie, Tian; Ravensbergen, Kristina; Hahm, Jong-in

    2016-02-14

    With the recent development of biomaterials and biodevices with reduced dimensionality, it is critical to comprehend protein adhesion processes to nanoscale solid surfaces, especially those occurring in a competitive adsorption environment. Complex sequences of adhesion events in competitive adsorption involving multicomponent protein systems have been extensively investigated, but our understanding is still limited primarily to macroscopic adhesion onto chemically simple surfaces. We examine the competitive adsorption behavior from a binary protein mixture containing bovine serum albumin and fibrinogen at the single protein level. We subsequently evaluate a series of adsorption and displacement processes occurring on both the macroscopic homopolymer and nanoscopic diblock copolymer surfaces, while systematically varying the protein concentration and incubation time. We identify the similarities and dissimilarities in competitive protein adsorption behavior between the two polymeric surfaces, the former presenting chemical uniformity at macroscale versus the latter exhibiting periodic nanointerfaces of chemically alternating polymeric segments. We then present our novel experimental finding of a large increase in the nanointerface-engaged residence time of the initially bound proteins and further explain the origin of this phenomenon manifested on nanoscale diblock copolymer surfaces. The outcomes of this study may provide timely insight into nanoscale competitive protein adsorption that is much needed in designing bioimplant and tissue engineering materials. In addition, the fundamental understanding gained from this study can be beneficial for the development of highly miniaturized biodevices and biomaterials fabricated by using nanoscale polymeric materials and interfaces.

  2. Investigation of protein-fatty acid interactions in zein films

    NASA Astrophysics Data System (ADS)

    Wang, Qin

    Zein, the prolamin of corn, has shown potential as industrial biopolymer for packaging and agricultural uses. Previous researchers plasticized zein with oleic acid and extruded it into sheets and films. Such products showed reasonable tensile and water barrier properties. However, those properties were affected by film structure, which is believed controlled by the interaction between zein and plasticizer. The nature of those interactions is still not well understood. Protein-fatty acid interactions in zein resin films were investigated by surface plasmon resonance and atomic force microscopy in this work. Preliminary research was conducted to establish the effect of solvent systems on the plasticization of zein by oleic acid. Although slight differences were observed, it was concluded that the use of 75% ethanol or 75% 2-propanol produced similar results. Surface plasmon resonance was employed to investigate zein static and dynamic adsorption on surfaces of hydrophilic and hydrophobic self-assembled monolayers (SAMs) generated by 11-mercaptoundecanoic acid or 1-octanethiol, respectively, and representing the two end groups of oleic acid. Results indicated that zein was adsorbed to both surfaces but showed higher affinity for hydrophilic groups. The corresponding thickness of zein specific binding layer on hydrophilic and hydrophobic SAMs was around 4.7 nm and 4.6 nm. Zein exhibited higher affinity for hydrophilic than for hydrophobic SAMs evidenced from the higher initial adsorption rate and ultimate surface coverage at all zein concentrations. Flushing surface with buffer would leave an apparent monolayer of zein, which is 5 times higher for hydrophilic than hydrophobic SAMs. This observation suggested that zein may use different sides of its molecule to interact with hydrophobic or hydrophilic groups. The surface topography of zein deposits on both SAMs was examined by atomic force microscopy. It was found that zein formed distinct ring-shaped structures with

  3. Unsaturated fatty acids supplementation reduces blood lead level in rats.

    PubMed

    Skoczyńska, Anna; Wojakowska, Anna; Nowacki, Dorian; Bobak, Łukasz; Turczyn, Barbara; Smyk, Beata; Szuba, Andrzej; Trziszka, Tadeusz

    2015-01-01

    Some dietary factors could inhibit lead toxicity. The aim of this study was to evaluate the effect of dietary compounds rich in unsaturated fatty acids (FA) on blood lead level, lipid metabolism, and vascular reactivity in rats. Serum metallothionein and organs' lead level were evaluated with the aim of assessing the possible mechanism of unsaturated FA impact on blood lead level. For three months, male Wistar rats that were receiving drinking water with (100 ppm Pb) or without lead acetate were supplemented per os daily with virgin olive oil or linseed oil (0.2 mL/kg b.w.) or egg derived lecithin fraction: "super lecithin" (50 g/kg b.w.). Mesenteric artery was stimulated ex vivo by norepinephrine (NE) administered at six different doses. Lecithin supplementation slightly reduced pressor responses of artery to NE. Lead administered to rats attenuated the beneficial effect of unsaturated FA on lipid metabolism and vascular reactivity to adrenergic stimulation. On the other hand, the super lecithin and linseed oil that were characterized by low omega-6 to omega-3 ratio (about 1) reduced the blood lead concentration. This effect was observed in lead poisoned rats (p < 0.0001) and also in rats nonpoisoned with lead (p < 0.05). PMID:26075218

  4. Unsaturated Fatty Acids Supplementation Reduces Blood Lead Level in Rats

    PubMed Central

    Skoczyńska, Anna; Wojakowska, Anna; Nowacki, Dorian; Bobak, Łukasz; Turczyn, Barbara; Smyk, Beata; Szuba, Andrzej; Trziszka, Tadeusz

    2015-01-01

    Some dietary factors could inhibit lead toxicity. The aim of this study was to evaluate the effect of dietary compounds rich in unsaturated fatty acids (FA) on blood lead level, lipid metabolism, and vascular reactivity in rats. Serum metallothionein and organs' lead level were evaluated with the aim of assessing the possible mechanism of unsaturated FA impact on blood lead level. For three months, male Wistar rats that were receiving drinking water with (100 ppm Pb) or without lead acetate were supplemented per os daily with virgin olive oil or linseed oil (0.2 mL/kg b.w.) or egg derived lecithin fraction: “super lecithin” (50 g/kg b.w.). Mesenteric artery was stimulated ex vivo by norepinephrine (NE) administered at six different doses. Lecithin supplementation slightly reduced pressor responses of artery to NE. Lead administered to rats attenuated the beneficial effect of unsaturated FA on lipid metabolism and vascular reactivity to adrenergic stimulation. On the other hand, the super lecithin and linseed oil that were characterized by low omega-6 to omega-3 ratio (about 1) reduced the blood lead concentration. This effect was observed in lead poisoned rats (p < 0.0001) and also in rats nonpoisoned with lead (p < 0.05). PMID:26075218

  5. Japanese encephalitis virus nonstructural protein NS5 interacts with mitochondrial trifunctional protein and impairs fatty acid β-oxidation.

    PubMed

    Kao, Yu-Ting; Chang, Bi-Lan; Liang, Jian-Jong; Tsai, Hang-Jen; Lee, Yi-Ling; Lin, Ren-Jye; Lin, Yi-Ling

    2015-03-01

    Infection with Japanese encephalitis virus (JEV) can induce the expression of pro-inflammatory cytokines and cause acute encephalitis in humans. β-oxidation breaks down fatty acids for ATP production in mitochondria, and impaired β-oxidation can induce pro-inflammatory cytokine expression. To address the role of fatty-acid β-oxidation in JEV infection, we measured the oxygen consumption rate of mock- and JEV-infected cells cultured with or without long chain fatty acid (LCFA) palmitate. Cells with JEV infection showed impaired LCFA β-oxidation and increased interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) expression. JEV nonstructural protein 5 (NS5) interacted with hydroxyacyl-CoA dehydrogenase α and β subunits, two components of the mitochondrial trifunctional protein (MTP) involved in LCFA β-oxidation, and NS5 proteins were detected in mitochondria and co-localized with MTP. LCFA β-oxidation was impaired and higher cytokines were induced in cells overexpressing NS5 protein as compared with control cells. Deletion and mutation studies showed that the N-terminus of NS5 was involved in the MTP association, and a single point mutation of NS5 residue 19 from methionine to alanine (NS5-M19A) reduced its binding ability with MTP. The recombinant JEV with NS5-M19A mutation (JEV-NS5-M19A) was less able to block LCFA β-oxidation and induced lower levels of IL-6 and TNF-α than wild-type JEV. Moreover, mice challenged with JEV-NS5-M19A showed less neurovirulence and neuroinvasiveness. We identified a novel function of JEV NS5 in viral pathogenesis by impairing LCFA β-oxidation and inducing cytokine expression by association with MTP.

  6. Japanese Encephalitis Virus Nonstructural Protein NS5 Interacts with Mitochondrial Trifunctional Protein and Impairs Fatty Acid β-Oxidation

    PubMed Central

    Kao, Yu-Ting; Chang, Bi-Lan; Liang, Jian-Jong; Tsai, Hang-Jen; Lee, Yi-Ling; Lin, Ren-Jye; Lin, Yi-Ling

    2015-01-01

    Infection with Japanese encephalitis virus (JEV) can induce the expression of pro-inflammatory cytokines and cause acute encephalitis in humans. β-oxidation breaks down fatty acids for ATP production in mitochondria, and impaired β-oxidation can induce pro-inflammatory cytokine expression. To address the role of fatty-acid β-oxidation in JEV infection, we measured the oxygen consumption rate of mock- and JEV-infected cells cultured with or without long chain fatty acid (LCFA) palmitate. Cells with JEV infection showed impaired LCFA β-oxidation and increased interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) expression. JEV nonstructural protein 5 (NS5) interacted with hydroxyacyl-CoA dehydrogenase α and β subunits, two components of the mitochondrial trifunctional protein (MTP) involved in LCFA β-oxidation, and NS5 proteins were detected in mitochondria and co-localized with MTP. LCFA β-oxidation was impaired and higher cytokines were induced in cells overexpressing NS5 protein as compared with control cells. Deletion and mutation studies showed that the N-terminus of NS5 was involved in the MTP association, and a single point mutation of NS5 residue 19 from methionine to alanine (NS5-M19A) reduced its binding ability with MTP. The recombinant JEV with NS5-M19A mutation (JEV-NS5-M19A) was less able to block LCFA β-oxidation and induced lower levels of IL-6 and TNF-α than wild-type JEV. Moreover, mice challenged with JEV-NS5-M19A showed less neurovirulence and neuroinvasiveness. We identified a novel function of JEV NS5 in viral pathogenesis by impairing LCFA β-oxidation and inducing cytokine expression by association with MTP. PMID:25816318

  7. Prediction of protein-protein interactions with clustered amino acids and weighted sparse representation.

    PubMed

    Huang, Qiaoying; You, Zhuhong; Zhang, Xiaofeng; Zhou, Yong

    2015-01-01

    With the completion of the Human Genome Project, bioscience has entered into the era of the genome and proteome. Therefore, protein-protein interactions (PPIs) research is becoming more and more important. Life activities and the protein-protein interactions are inseparable, such as DNA synthesis, gene transcription activation, protein translation, etc. Though many methods based on biological experiments and machine learning have been proposed, they all spent a long time to learn and obtained an imprecise accuracy. How to efficiently and accurately predict PPIs is still a big challenge. To take up such a challenge, we developed a new predictor by incorporating the reduced amino acid alphabet (RAAA) information into the general form of pseudo-amino acid composition (PseAAC) and with the weighted sparse representation-based classification (WSRC). The remarkable advantages of introducing the reduced amino acid alphabet is being able to avoid the notorious dimensionality disaster or overfitting problem in statistical prediction. Additionally, experiments have proven that our method achieved good performance in both a low- and high-dimensional feature space. Among all of the experiments performed on the PPIs data of Saccharomyces cerevisiae, the best one achieved 90.91% accuracy, 94.17% sensitivity, 87.22% precision and a 83.43% Matthews correlation coefficient (MCC) value. In order to evaluate the prediction ability of our method, extensive experiments are performed to compare with the state-of-the-art technique, support vector machine (SVM). The achieved results show that the proposed approach is very promising for predicting PPIs, and it can be a helpful supplement for PPIs prediction. PMID:25984606

  8. Analysis of the interactions of sulfur-containing amino acids in membrane proteins.

    PubMed

    Gómez-Tamayo, José C; Cordomí, Arnau; Olivella, Mireia; Mayol, Eduardo; Fourmy, Daniel; Pardo, Leonardo

    2016-08-01

    The interactions of Met and Cys with other amino acid side chains have received little attention, in contrast to aromatic-aromatic, aromatic-aliphatic or/and aliphatic-aliphatic interactions. Precisely, these are the only amino acids that contain a sulfur atom, which is highly polarizable and, thus, likely to participate in strong Van der Waals interactions. Analysis of the interactions present in membrane protein crystal structures, together with the characterization of their strength in small-molecule model systems at the ab-initio level, predicts that Met-Met interactions are stronger than Met-Cys ≈ Met-Phe ≈ Cys-Phe interactions, stronger than Phe-Phe ≈ Phe-Leu interactions, stronger than the Met-Leu interaction, and stronger than Leu-Leu ≈ Cys-Leu interactions. These results show that sulfur-containing amino acids form stronger interactions than aromatic or aliphatic amino acids. Thus, these amino acids may provide additional driving forces for maintaining the 3D structure of membrane proteins and may provide functional specificity.

  9. Regulation of collagenase gene expression by okadaic acid, an inhibitor of protein phosphatases.

    PubMed Central

    Kim, S J; Lafyatis, R; Kim, K Y; Angel, P; Fujiki, H; Karin, M; Sporn, M B; Roberts, A B

    1990-01-01

    Human collagenase gene expression is regulated transcriptionally and is inducible by various mitogens in many cell types. To investigate the molecular mechanisms of this response, we examined the effects on collagenase gene expression of okadaic acid, a non-12-O-tetradecanoyl-phorbol-13-acetate (TPA)-type tumor promoter, which induces apparent "activation" of protein kinases by inhibition of protein phosphatases. Steady state levels of collagenase mRNA were markedly increased by okadaic acid treatment. We show that the AP-1 consensus sequence in the collagenase promoter is required for the induction of collagenase gene expression by okadaic acid, even though sequences upstream of the AP-1 consensus site have an additive effect. We also examined the regulation by okadaic acid of expression of the components of the AP-1 complex, c-fos and c-jun. c-fos expression is dramatically stimulated by okadaic acid, whereas c-jun expression is stimulated to a lesser extent. Induction of c-fos gene mRNA occurs through a region known to contain multiple regulatory elements. These results suggest that phosphorylation regulates collagenase gene expression mediated by an AP-1 binding site. Images PMID:1966042

  10. Proximate composition, fatty acid analysis and protein digestibility-corrected amino acid score of three Mediterranean cephalopods.

    PubMed

    Zlatanos, Spiros; Laskaridis, Kostas; Feist, Christian; Sagredos, Angelos

    2006-10-01

    Proximate composition, fatty acid analysis and protein digestibility-corrected amino acid score (PDCAAS) in three commercially important cephalopods of the Mediterranean sea (cuttlefish, octopus and squid) were determined. The results of the proximate analysis showed that these species had very high protein:fat ratios similar to lean beef. Docosahexaenoic, palmitic and eicosipentaenoic acid were the most abundant fatty acids among analyzed species. The amount of n-3 fatty acids was higher than that of saturated, monounsaturated and n-6 fatty acids. Despite the fact that cephalopods contain small amounts of fat they were found quite rich in n-3 fatty acids. Finally, PDCAAS indicated that these organisms had a very good protein quality.

  11. Low irradiances affect abscisic acid, indole-3-acidic acid, and cytokinin levels of wheat (Triticum aestivum L.) tissues

    NASA Technical Reports Server (NTRS)

    Nan, R.; Carman, J. G.; Salisbury, F. B.

    1999-01-01

    Wheat (Triticum aestivum L.) plants were grown under four irradiance levels: 1,400, 400, 200, and 100 micromol m-2 s-1. Leaves and roots were sampled before, during, and after the boot stage, and levels of abscisic acid (ABA), indole-3-acetic acid (IAA), zeatin, zeatin riboside, dihydrozeatin, dihydrozeatin riboside, isopentenyl adenine, and isopentenyl adenosine were quantified using noncompetitive indirect ELISA systems. Levels of IAA in leaves and roots of plants exposed to 100 micromol m-2 s-1 of irradiance were 0.7 and 2.9 micromol kg-1 dry mass (DM), respectively. These levels were 0.2 and 1.0 micromol kg-1 DM, respectively, when plants were exposed to 1,400 micromol m-2 s-1. Levels of ABA in leaves and roots of plants exposed to 100 micromol m-2 s-1 were 0.65 and 0.55 micromol kg-1 DM, respectively. They were 0.24 micromol kg-1 DM (both leaves and roots) when plants were exposed to 1,400 micromol m-2 s-1. Levels of isopentenyl adenosine in leaves (24.3 nmol kg-1 DM) and roots (29.9 nmol kg-1 DM) were not affected by differences in the irradiance regime. Similar values were obtained in a second experiment. Other cytokinins could not be detected (<10 nmol kg 1 DM) in either experiment with the sample sizes used (150-600 mg DM for roots and shoots, respectively).

  12. Mitogen-activated protein kinase and abscisic acid signal transduction.

    PubMed

    Heimovaara-Dijkstra, S; Testerink, C; Wang, M

    2000-01-01

    The phytohormone abscisic acid (ABA) is a classical plant hormone, responsible for regulation of abscission, diverse aspects of plant and seed development, stress responses and germination. It was found that ABA signal transduction in plants can involve the activity of type 2C-phosphatases (PP2C), calcium, potassium, pH and a transient activation of MAP kinase. The ABA signal transduction cascades have been shown to be tissue-specific, the transient activation of MAP kinase has until now only been found in barley aleurone cells. However, type 2C phosphatases are involved in the induction of most ABA responses, as shown by the PP2C-deficient abi-mutants. These phosphatases show high homology with phosphatases that regulate MAP kinase activity in yeast. In addition, the role of farnesyl transferase as a negative regulator of ABA responses also indicates towards involvement of MAP kinase in ABA signal transduction. Farnesyl transferase is known to regulate Ras proteins, Ras proteins in turn are known to regulate MAP kinase activation. Interestingly, Ras-like proteins were detected in barley aleurone cells. Further establishment of the involvement of MAP kinase in ABA signal transduction and its role therein, still awaits more study.

  13. Nucleic Acids for Ultra-Sensitive Protein Detection

    PubMed Central

    Janssen, Kris P. F.; Knez, Karel; Spasic, Dragana; Lammertyn, Jeroen

    2013-01-01

    Major advancements in molecular biology and clinical diagnostics cannot be brought about strictly through the use of genomics based methods. Improved methods for protein detection and proteomic screening are an absolute necessity to complement to wealth of information offered by novel, high-throughput sequencing technologies. Only then will it be possible to advance insights into clinical processes and to characterize the importance of specific protein biomarkers for disease detection or the realization of “personalized medicine”. Currently however, large-scale proteomic information is still not as easily obtained as its genomic counterpart, mainly because traditional antibody-based technologies struggle to meet the stringent sensitivity and throughput requirements that are required whereas mass-spectrometry based methods might be burdened by significant costs involved. However, recent years have seen the development of new biodetection strategies linking nucleic acids with existing antibody technology or replacing antibodies with oligonucleotide recognition elements altogether. These advancements have unlocked many new strategies to lower detection limits and dramatically increase throughput of protein detection assays. In this review, an overview of these new strategies will be given. PMID:23337338

  14. Inhibition of the Hematopoietic Protein Tyrosine Phosphatase by Phenoxyacetic Acids.

    PubMed

    Bobkova, Ekaterina V; Liu, Wallace H; Colayco, Sharon; Rascon, Justin; Vasile, Stefan; Gasior, Carlton; Critton, David A; Chan, Xochella; Dahl, Russell; Su, Ying; Sergienko, Eduard; Chung, Thomas D Y; Mustelin, Tomas; Page, Rebecca; Tautz, Lutz

    2011-02-01

    Protein tyrosine phosphatases (PTPs) have only recently become the focus of attention in the search for novel drug targets despite the fact that they play vital roles in numerous cellular processes and are implicated in many human diseases. The hematopoietic protein tyrosine phosphatase (HePTP) is often found dysregulated in preleukemic myelodysplastic syndrome (MDS), as well as in acute myelogenous leukemia (AML). Physiological substrates of HePTP include the mitogen-activated protein kinases (MAPKs) ERK1/2 and p38. Specific modulators of HePTP catalytic activity will be useful for elucidating mechanisms of MAPK regulation in hematopietic cells, and may also provide treatments for hematopoietic malignancies such as AML. Here we report the discovery of phenoxyacetic acids as inhibitors of HePTP. Structure-activity relationship (SAR) analysis and in silico docking studies reveal the molecular basis of HePTP inhibition by these compounds. We also show that these compounds are able to penetrate cell membranes and inhibit HePTP in human T lymphocytes.

  15. Ultrasensitive detection of proteins and sugars at single-cell level

    PubMed Central

    Watabe, Satoshi; Morikawa, Mika; Kaneda, Mugiho; Nakaishi, Kazunari; Nakatsuma, Akira; Ninomiya, Masaki; Yoshimura, Teruki; Miura, Toshiaki; Ito, Etsuro

    2016-01-01

    ABSTRACT Each cell produces its own responses even if it appears identical to other cells. To analyze these individual cell characteristics, we need to measure trace amounts of molecules in a single cell. Nucleic acids in a single cell can be easily amplified by polymerase chain reaction, but single-cell measurement of proteins and sugars will require de novo techniques. In the present study, we outline the techniques we have developed toward this end. For proteins, our ultrasensitive enzyme-linked immunosorbent assay (ELISA) coupled with thionicotinamide-adenine dinucleotide cycling can detect proteins at subattomoles per assay. For sugars, fluorescence correlation spectroscopy coupled with glucose oxidase-catalyzed reaction allows us to measure glucose at tens of nM. Our methods thus offer versatile techniques for single-cell-level analyses, and they are hoped to strongly promote single-cell biology as well as to develop noninvasive tests in clinical medicine. PMID:27064305

  16. An efficient protocol for incorporation of an unnatural amino acid in perdeuterated recombinant proteins using glucose-based media.

    PubMed

    Venditti, Vincenzo; Fawzi, Nicolas L; Clore, G Marius

    2012-03-01

    The in vivo incorporation of unnatural amino acids into proteins is a well-established technique requiring an orthogonal tRNA/aminoacyl-tRNA synthetase pair specific for the unnatural amino acid that is incorporated at a position encoded by a TAG amber codon. Although this technology provides unique opportunities to engineer protein structures, poor protein yields are usually obtained in deuterated media, hampering its application in the protein NMR field. Here, we describe a novel protocol for incorporating unnatural amino acids into fully deuterated proteins using glucose-based media (which are relevant to the production, for example, of amino acid-specific methyl-labeled proteins used in the study of large molecular weight systems). The method consists of pre-induction of the pEVOL plasmid encoding the tRNA/aminoacyl-tRNA synthetase pair in a rich, H(2)O-based medium prior to exchanging the culture into a D(2)O-based medium. Our protocol results in high level of isotopic incorporation (~95%) and retains the high expression level of the target protein observed in Luria-Bertani medium. PMID:22350951

  17. Codon-level information improves predictions of inter-residue contacts in proteins by correlated mutation analysis.

    PubMed

    Jacob, Etai; Unger, Ron; Horovitz, Amnon

    2015-09-15

    Methods for analysing correlated mutations in proteins are becoming an increasingly powerful tool for predicting contacts within and between proteins. Nevertheless, limitations remain due to the requirement for large multiple sequence alignments (MSA) and the fact that, in general, only the relatively small number of top-ranking predictions are reliable. To date, methods for analysing correlated mutations have relied exclusively on amino acid MSAs as inputs. Here, we describe a new approach for analysing correlated mutations that is based on combined analysis of amino acid and codon MSAs. We show that a direct contact is more likely to be present when the correlation between the positions is strong at the amino acid level but weak at the codon level. The performance of different methods for analysing correlated mutations in predicting contacts is shown to be enhanced significantly when amino acid and codon data are combined.

  18. Codon-level information improves predictions of inter-residue contacts in proteins by correlated mutation analysis

    PubMed Central

    Jacob, Etai; Unger, Ron; Horovitz, Amnon

    2015-01-01

    Methods for analysing correlated mutations in proteins are becoming an increasingly powerful tool for predicting contacts within and between proteins. Nevertheless, limitations remain due to the requirement for large multiple sequence alignments (MSA) and the fact that, in general, only the relatively small number of top-ranking predictions are reliable. To date, methods for analysing correlated mutations have relied exclusively on amino acid MSAs as inputs. Here, we describe a new approach for analysing correlated mutations that is based on combined analysis of amino acid and codon MSAs. We show that a direct contact is more likely to be present when the correlation between the positions is strong at the amino acid level but weak at the codon level. The performance of different methods for analysing correlated mutations in predicting contacts is shown to be enhanced significantly when amino acid and codon data are combined. DOI: http://dx.doi.org/10.7554/eLife.08932.001 PMID:26371555

  19. Contribution of polyunsaturated fatty acids to intestinal repair in protein-energy malnutrition.

    PubMed

    Nieto, Natalia; Mesa, María Dolores; López-Pedrosa, José María; Torres, M Isabel; Ríos, Antonio; Suárez, María Dolores; Gil, Angel

    2007-06-01

    The aim of this study was to assess the effect of polyunsaturated fatty acids supplied in the diet on intestinal mucosa repair in a rat model of protein-energy malnutrition. Rats were fed either a standard semipurified diet or the same diet containing lactose as the only source of carbohydrate to cause protein-energy malnutrition. Diarrhea was induced within 24 h and was maintained for 2 weeks, after which both groups of rats were fed for 1 week either the standard diet or the standard diet supplemented with different sources of fatty acids, such as olive oil (OO), fish oil (FO), and purified phospholipids from pig brain (BPL). The lactose-enriched diet caused loss of enterocyte microvilli, lymphocyte infiltration, supranuclear cytoplasmic vesiculation, decreased number of goblet cells, low-density enlarged mitochondria, and less cristae. The FO diet improved the pathology score according to the histological and ultrastructural analysis, with an increased number of goblet cells, ratio of microvilli length to crypt depth, and percentage of intraepithelial lymphocytes compared to those found in rats with protein-energy malnutrition. We previously reported that chronic diarrhea depletes the antioxidant defense in rat intestine; we now show that both, the FO and the BPL diets, increase GSH levels in colon and that some antioxidant enzyme activities vary according to the source of fatty acids, with higher catalase and superoxide dismutase by the FO diet in jejunum, increased catalase by the BPL diet in jejunum, and elevated glutathione peroxidase by the OO diet in colon. The fatty acid profile of intestinal mucosa reflects the source of fat in the diet, with the lowest ratio of n-6/n-3 for rats fed the FO diet. These results suggest that dietary polyunsaturated fatty acids, particularly those in the n-3 series, may play an important role in intestinal repair in chronic diarrhea due to protein-energy malnutrition.

  20. Computationally Efficient Multiscale Reactive Molecular Dynamics to Describe Amino Acid Deprotonation in Proteins

    PubMed Central

    2016-01-01

    An important challenge in the simulation of biomolecular systems is a quantitative description of the protonation and deprotonation process of amino acid residues. Despite the seeming simplicity of adding or removing a positively charged hydrogen nucleus, simulating the actual protonation/deprotonation process is inherently difficult. It requires both the explicit treatment of the excess proton, including its charge defect delocalization and Grotthuss shuttling through inhomogeneous moieties (water and amino residues), and extensive sampling of coupled condensed phase motions. In a recent paper (J. Chem. Theory Comput.2014, 10, 2729−273725061442), a multiscale approach was developed to map high-level quantum mechanics/molecular mechanics (QM/MM) data into a multiscale reactive molecular dynamics (MS-RMD) model in order to describe amino acid deprotonation in bulk water. In this article, we extend the fitting approach (called FitRMD) to create MS-RMD models for ionizable amino acids within proteins. The resulting models are shown to faithfully reproduce the free energy profiles of the reference QM/MM Hamiltonian for PT inside an example protein, the ClC-ec1 H+/Cl– antiporter. Moreover, we show that the resulting MS-RMD models are computationally efficient enough to then characterize more complex 2-dimensional free energy surfaces due to slow degrees of freedom such as water hydration of internal protein cavities that can be inherently coupled to the excess proton charge translocation. The FitRMD method is thus shown to be an effective way to map ab initio level accuracy into a much more computationally efficient reactive MD method in order to explicitly simulate and quantitatively describe amino acid protonation/deprotonation in proteins. PMID:26734942

  1. Relation of soya bean meal level to the concentration of plasma free amino acids and body growth in white rats.

    PubMed

    Mandal, Tapas K; Parvin, Nargish; Mondal, Santanu; Saxena, Vijaylaxmi; Saxena, Ashok K; Sarkar, Sabyasachi; Saha, Mitali

    2012-04-01

    Amino acid (AA) levels in plasma and body growth were determined in rats (n20) fed diets with different soya bean meal levels. Free AA in plasma was determined by reversed-phase high-pressure liquid chromatography. We have used four levels of protein diets like 8%, 15%, 23% and 35% in this trial. Rats which were fed the low-protein (8%) diet with low percentage of soya bean meal were found to be growth-retarded. The body weight gain of high protein group (35%) was lower than that of the 23% groups. In the rats fed with the low-soya bean meal diet, some nonessential AA (NEAA) in plasma like asparagine, aspartic acid, cysteine, glutamic acid and serine increased, whereas the essential AA (EAA), with the exception of arginine, methionine and valine decreased. Here, plasma EAA-to-NEAA ratios were not correlated to growth and experimental diet. We hypothesize that AA metabolism is associated to changes in growth in rats on different protein intake. This study has showed the sensitivity of body mass gain, feed intake, feed conversion rate of rats to four levels of protein in the diet under controlled experimental conditions.

  2. Bone morphogenetic protein (BMP) signaling regulates mitotic checkpoint protein levels in human breast cancer cells.

    PubMed

    Yan, Hualong; Zhu, Songcheng; Song, Chenlin; Liu, Naifa; Kang, Jiuhong

    2012-04-01

    Aberrant expression of mitotic checkpoint genes compromises mitotic checkpoint, leads to chromosome instability and tumorigenesis. However, the cell signals that control mitotic checkpoint gene expression have not been reported so far. In the present study we show that, in human breast cancer cells, chemical inhibition of Bone morphogenetic proteins (BMPs), but not Transforming Growth Factor-β (TGF-β), abrogates the mitotic arrest induced by nocodazole. Protein expression analysis reveals that inhibition of BMP signaling dramatically down regulates protein levels of mitotic checkpoint components BUB3, Hec1, TTK and MAD2, but inhibition of TGF-β has relatively minor effect on the expression of these proteins. Activation of BMP signaling specifically up regulates BUB3, and activation of Activin A signaling globally down regulates these proteins level. Furthermore, overexpressing MAD2, TTK, BUB3 or Hec1 significantly rescues the mitotic arrest defect caused by BMP inhibition. Our results demonstrated for the first time that TGF-β family cytokines are cellular signals regulating mitotic checkpoint and perturbations in intrinsic BMP signaling could lead to suppression of mitotic checkpoint signaling by downregulating key checkpoint proteins. The results suggest a possible mechanism by which dysregulation of TGF-β signaling causes mitotic checkpoint defects and drives tumorigenesis. The finding also provides a potential and more specific strategy for cancer prevention by targeting BMP and mitotic checkpoint connection. PMID:22234345

  3. Defining meal requirements for protein to optimize metabolic roles of amino acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dietary protein provides essential amino acids (EAAs) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to postprandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signal...

  4. Defining meal requirements for protein to optimize metabolic roles of amino acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dietary protein provides essential amino acids (EAA) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to post-prandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signal...

  5. Limiting amino acid for protein synthesis with mammary cells in tissue culture.

    PubMed

    Park, C S; Chandler, P T; Norman, A W

    1976-05-01

    To identify the limiting amino acid in the minimal essential medium as published by Eagle (Science 130:432, 1959) for milk protein synthesis in rat mammary cells in tissue culture, two different experimental approaches were used. The first study involved the reduction of amino acids singly from the total amino acid complement of the medium for milk protein synthesis. The second study was to investigate the effect on milk protein synthesis of single amino acid addition to the basic complement of amino acids. Order of limiting amino acids was lysine (first) and possible methionine, valine, or arginine (second).

  6. Alterations of Amino Acid Level in Depressed Rat Brain

    PubMed Central

    Yang, Pei; Li, Xuechun; Tian, Jingchen; Jing, Fu; Qu, Changhai; Lin, Longfei; Zhang, Hui

    2014-01-01

    Amino-acid neurotransmitter system dysfunction plays a major role in the pathophysiology of depression. Several studies have demonstrated the potential of amino acids as a source of neuro-specific biomarkers could be used in future diagnosis of depression. Only partial amino acids such as glycine and asparagine were determined from certain parts of rats' brain included hippocampi and cerebral cortex in previous studies. However, according to systematic biology, amino acids in different area of brain are interacted and interrelated. Hence, the determination of 34 amino acids through entire rats' brain was conducted in this study in order to demonstrate more possibilities for biomarkers of depression by discovering other potential amino acids in more areas of rats' brain. As a result, 4 amino acids (L-aspartic acid, L-glutamine, taurine and γ-amino-n-butyric acid) among 34 were typically identified as potentially primary biomarkers of depression by data statistics. Meanwhile, an antidepressant called Fluoxetine was employed to verify other potential amino acids which were not identified by data statistics. Eventually, we found L-α-amino-adipic acid could also become a new potentially secondary biomarker of depression after drug validation. In conclusion, we suggested that L-aspartic acid, L-glutamine, taurine, γ-amino-n-butyric acid and L-α-amino-adipic acid might become potential biomarkers for future diagnosis of depression and development of antidepressant. PMID:25352755

  7. Cochlear ablation effects on amino acid levels in the chinchilla cochlear nucleus.

    PubMed

    Godfrey, D A; Chen, K; Godfrey, M A; Lee, A C; Crass, S P; Shipp, D; Simo, H; Robinson, K T

    2015-06-25

    Inner ear damage can lead to hearing disorders, including tinnitus, hyperacusis, and hearing loss. We measured the effects of severe inner ear damage, produced by cochlear ablation, on the levels and distributions of amino acids in the first brain center of the auditory system, the cochlear nucleus. Measurements were also made for its projection pathways and the superior olivary nuclei. Cochlear ablation produces complete degeneration of the auditory nerve, which provides a baseline for interpreting the effects of partial damage to the inner ear, such as that from ototoxic drugs or intense sound. Amino acids play a critical role in neural function, including neurotransmission, neuromodulation, cellular metabolism, and protein construction. They include major neurotransmitters of the brain - glutamate, glycine, and γ-aminobutyrate (GABA) - as well as others closely related to their metabolism and/or functions - aspartate, glutamine, and taurine. Since the effects of inner ear damage develop over time, we measured the changes in amino acid levels at various survival times after cochlear ablation. Glutamate and aspartate levels decreased by 2weeks in the ipsilateral ventral cochlear nucleus and deep layer of the dorsal cochlear nucleus, with the largest decreases in the posteroventral cochlear nucleus (PVCN): 66% for glutamate and 63% for aspartate. Aspartate levels also decreased in the lateral part of the ipsilateral trapezoid body, by as much as 50%, suggesting a transneuronal effect. GABA and glycine levels showed some bilateral decreases, especially in the PVCN. These results may represent the state of amino acid metabolism in the cochlear nucleus of humans after removal of eighth nerve tumors, which may adversely result in destruction of the auditory nerve. Measurement of chemical changes following inner ear damage may increase understanding of the pathogenesis of hearing impairments and enable improvements in their diagnosis and treatment. PMID:25839146

  8. Site-specific incorporation of redox active amino acids into proteins

    DOEpatents

    Alfonta, Lital; Schultz, Peter G.; Zhang, Zhiwen

    2009-02-24

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  9. Site-specific incorporation of redox active amino acids into proteins

    DOEpatents

    Alfonta; Lital , Schultz; Peter G. , Zhang; Zhiwen

    2010-10-12

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  10. Site-specific incorporation of redox active amino acids into proteins

    SciTech Connect

    Alfonta, Lital; Schultz, Peter G.; Zhang, Zhiwen

    2012-02-14

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  11. Site-specific incorporation of redox active amino acids into proteins

    SciTech Connect

    Alfonta, Lital; Schultz, Peter G.; Zhang, Zhiwen

    2011-08-30

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  12. Serum uric acid levels during leprosy reaction episodes

    PubMed Central

    Alves-Junior, Eduardo R.; Arruda, Talita A.; Lopes, Jose C.; Fontes, Cor J.F.

    2016-01-01

    Background. Leprosy reactions are acute inflammatory episodes that occur mainly in the multibacillary forms of the disease. The reactions are classified as type 1 (reverse reaction) or type 2 (erythema nodosum leprosum). Leprosy-associated oxidative stress has been widely demonstrated. Several recent studies have shown uric acid (UA) to have antioxidative effects under pathologic conditions. The objective of this study was to assess serum levels of UA in patients with leprosy reactions, with the aim of monitoring their levels before and after treatment, compared with levels in leprosy patients without reactions. Methods. The study included patients aged 18–69 years assisted at a leprosy treatment reference center in the Central Region of Brazil. Patients who were pregnant; were using immunosuppressant drugs or immunobiologicals; or had an autoimmune disease, human immunodeficiency virus infection, acquired immune deficiency syndrome, or tuberculosis were excluded. Upon recruitment, all individuals were clinically assessed for skin lesions and neural or systemic impairment. Some patients had already completed treatment for leprosy, while others were still undergoing treatment or had initiated treatment after being admitted. The treatment of the reactional episode was started only after the initial evaluation. Laboratory assessments were performed upon admission (baseline) and at approximately 30 and 60 days (time points 1 and 2, respectively). Results. A total of 123 leprosy patients were recruited between June 2012 and June 2015; among them, 56, 42, and 25 presented with type 1, type 2, and no reactions, respectively. Serum UA levels were significantly reduced in patients with type 2 leprosy reactions compared with patients in the control group and remained lower in the two subsequent assessments, after initiation of anti-reaction treatments, with similar values to those recorded before the treatment. Discussion. The decreased serum UA levels in patients with

  13. Ascorbic acid levels of aqueous humor of dogs after experimental phacoemulsification.

    PubMed

    De Biaggi, Christianni P; Barros, Paulo S M; Silva, Vanessa V; Brooks, Dennis E; Barros, Silvia B M

    2006-01-01

    Phacoemulsification has been successfully employed in humans and animals for lens extraction. This ultrasonic extracapsular surgical technique induces hydroxyl radical formation in the anterior chamber, which accumulates despite irrigation and aspiration. In this paper we determined the total antioxidant status of aqueous humor after phacoemulsification by measuring aqueous humor ascorbic acid levels. Mixed-breed dogs (n = 11; weighing about 10 kg) with normal eyes as determined by slit-lamp biomicroscopy, applanation tonometry, and indirect ophthalmoscopy had phacoemulsification performed in one eye with the other eye used as a control. Samples of aqueous humor were obtained by anterior chamber paracentesis before surgery and at days 1, 2, 3, 7, and 15 after surgery. Total aqueous humor antioxidant status was inferred from the capacity of aqueous humor to inhibit free radical generation by 2,2-azobis (2-amidopropane) chlorine. Ascorbic acid concentrations were measured by high-pressure liquid chromatography with UV detection. Protein content was determined with the biuret reagent. Statistical analysis was performed by anova followed by the paired t-test. Total antioxidant capacity was reduced from 48 to 27 min during the first 24 h with a gradual increase thereafter, remaining statistically lower than the control eye until 7 days postoperatively. Reduced levels of ascorbic acid followed this reduction in antioxidant capacity (from 211 to 99 microm after 24 h), remaining lower than the control eye until 15 days postoperatively. Protein concentration in aqueous humor increased from 0.62 mg/mL to 30.8 mg/mL 24 h after surgery, remaining statistically lower than the control eye until 15 days postoperatively. Paracentesis alone did not significantly alter the parameters measured. These results indicate that after phacoemulsification, the aqueous humor ascorbic acid levels and antioxidant defenses in aqueous humor are reduced, indirectly corroborating free radical

  14. Interconnection between the protein solubility and amino acid and dipeptide compositions.

    PubMed

    Niu, Xiaohui; Li, Nana; Chen, Dinyan; Wang, Zengzhen

    2013-01-01

    Obtaining soluble proteins in sufficient concentrations helps increase the overall success rate in various experimental studies. Protein solubility is an individual trait ultimately determined by its primary protein sequence. Exploring the interconnection between the protein solubility and the compositions of protein sequence is instrumental for setting priorities on targets in large scale proteomics projects. In this paper, amino acid composition (20 dimensions) and the dipeptide composition (400 dimensions) were extracted to form the total candidate feature pool (420 dimensions), and each feature was selected into the feature vectors one by one, which were sorted by the absolute value of the correlation coefficient. Finally, we evaluated and recorded the 420 results of Support Vector Machine (SVM) as the prediction engine. According to the results of SVM, the first 208 features were chosen from the 420 dimensions, which were considered as the efficient ones. By analyzing the composition of the former 208 features, we found that the protein solubility was significantly influenced by the occurrence frequencies of the acidic amino acids, basic amino acids, non-polar hydrophobic amino acids and the two polar neutral amino acids(C, Q) in the protein sequences. Additionally, we detected that the dipeptides composed by the acidic amino acids (D, E) and basic amino acids (K, R and H), especially the dipeptide composed by the acidic amino acids (D, E), had strong interconnection with the protein solubility.

  15. Topology of RNA-protein nucleobase-amino acid π-π interactions and comparison to analogous DNA-protein π-π contacts.

    PubMed

    Wilson, Katie A; Holland, Devany J; Wetmore, Stacey D

    2016-05-01

    The present work analyzed 120 high-resolution X-ray crystal structures and identified 335 RNA-protein π-interactions (154 nonredundant) between a nucleobase and aromatic (W, H, F, or Y) or acyclic (R, E, or D) π-containing amino acid. Each contact was critically analyzed (including using a visual inspection protocol) to determine the most prevalent composition, structure, and strength of π-interactions at RNA-protein interfaces. These contacts most commonly involve F and U, with U:F interactions comprising one-fifth of the total number of contacts found. Furthermore, the RNA and protein π-systems adopt many different relative orientations, although there is a preference for more parallel (stacked) arrangements. Due to the variation in structure, the strength of the intermolecular forces between the RNA and protein components (as determined from accurate quantum chemical calculations) exhibits a significant range, with most of the contacts providing significant stability to the associated RNA-protein complex (up to -65 kJ mol(-1)). Comparison to the analogous DNA-protein π-interactions emphasizes differences in RNA- and DNA-protein π-interactions at the molecular level, including the greater abundance of RNA contacts and the involvement of different nucleobase/amino acid residues. Overall, our results provide a clearer picture of the molecular basis of nucleic acid-protein binding and underscore the important role of these contacts in biology, including the significant contribution of π-π interactions to the stability of nucleic acid-protein complexes. Nevertheless, more work is still needed in this area in order to further appreciate the properties and roles of RNA nucleobase-amino acid π-interactions in nature.

  16. Effects of conjugated linoleic acid supplementation and feeding level on dairy performance, milk fatty acid composition, and body fat changes in mid-lactation goats.

    PubMed

    Ghazal, S; Berthelot, V; Friggens, N C; Schmidely, P

    2014-11-01

    The objective of this trial was to study the interaction between the supplementation of lipid-encapsulated conjugated linoleic acid (CLA; 4.5 g of cis-9,trans-11 C18:2 and 4.5 g of trans-10,cis-12 C18:2) and feeding level to test if milk performance or milk fatty acid (FA) profile are affected by the interaction between CLA and feeding level. Twenty-four dairy goats were used in an 8-wk trial with a 3-wk adaptation to the experimental ration that contained corn silage, beet pulp, barley, and a commercial concentrate. During the third week, goats were assigned into blocks of 2 goats according to their dry matter intake (DMI), raw milk yield, and fat yield. Each block was randomly allocated to control (45 g of Ca salt of palm oil/d) or CLA treatment. Within each block, one goat was fed to cover 100% (FL100) of the calculated energy requirements and the other was fed 85% of the DMI of the first goat (FL85). Individual milk production and composition were recorded weekly, and milk FA composition was analyzed in wk 3, 5, and 7. Conjugated linoleic acid supplementation reduced milk fat content and fat yield by 17 and 19%, respectively, independent of the feeding level. It reduced both the secretion of milk FA synthesized de novo, and those taken up from the blood. No interaction between CLA and feeding level was observed on milk secretion of any group of FA. The CLA supplementation had no effect on DMI, milk yield, protein, and lactose yields but it improved calculated net energy for lactation balance. Goats fed the FL100 × CLA diet tended to have the highest DMI and protein yield. The interaction between CLA and feeding level was not significant for any other variables. Compared with the goats fed FL100, those fed FL85 had lower DMI, lower net energy for lactation balance, and lower digestible protein in the intestine balance. The body weight; milk yield; milk fat, protein, and lactose yields; and fat, protein, lactose, and urea contents in milk were not affected by

  17. Extraction of proteins with low fluoride level from Antarctic krill (Euphausia superba) and their composition analysis.

    PubMed

    Wang, Lingzhao; Xue, Changhu; Wang, Yuming; Yang, Bao

    2011-06-01

    The extraction of proteins with low fluoride level (LFP) from Antarctic krill (Euphausia superba) was investigated in this work. The optimal conditions for protein solubilization were determined to be pH 11.5 and 4 °C. The proteins were solubilized two times; a water/krill ratio (mL/g) of 6 and a time of 30 min were used for the first step, whereas the second used a water/krill residue ratio (mL/g) of 3 and a time of 30 min. The optimum pH for protein precipitation was 4.6. A LFP with fluoride content of 9.86 mg/kg (dry weight) was finally obtained through a fluoride removal program. The protein yield of LFP was 52.68%. Composition analysis of LFP indicated it was composed of 66.96% of crude proteins (dry weight) and 33.01% of total lipids (dry weight),, and all nine essential amino acids were in sufficient amounts to meet FAO/WHO/UNU requirements for adults and infants. In addition, LFP could be taken as a good source of EPA and DHA for consideration of use as a food item for human consumption.

  18. Ascertaining effects of nanoscale polymeric interfaces on competitive protein adsorption at the individual protein level

    NASA Astrophysics Data System (ADS)

    Song, Sheng; Xie, Tian; Ravensbergen, Kristina; Hahm, Jong-In

    2016-02-01

    With the recent development of biomaterials and biodevices with reduced dimensionality, it is critical to comprehend protein adhesion processes to nanoscale solid surfaces, especially those occurring in a competitive adsorption environment. Complex sequences of adhesion events in competitive adsorption involving multicomponent protein systems have been extensively investigated, but our understanding is still limited primarily to macroscopic adhesion onto chemically simple surfaces. We examine the competitive adsorption behavior from a binary protein mixture containing bovine serum albumin and fibrinogen at the single protein level. We subsequently evaluate a series of adsorption and displacement processes occurring on both the macroscopic homopolymer and nanoscopic diblock copolymer surfaces, while systematically varying the protein concentration and incubation time. We identify the similarities and dissimilarities in competitive protein adsorption behavior between the two polymeric surfaces, the former presenting chemical uniformity at macroscale versus the latter exhibiting periodic nanointerfaces of chemically alternating polymeric segments. We then present our novel experimental finding of a large increase in the nanointerface-engaged residence time of the initially bound proteins and further explain the origin of this phenomenon manifested on nanoscale diblock copolymer surfaces. The outcomes of this study may provide timely insight into nanoscale competitive protein adsorption that is much needed in designing bioimplant and tissue engineering materials. In addition, the fundamental understanding gained from this study can be beneficial for the development of highly miniaturized biodevices and biomaterials fabricated by using nanoscale polymeric materials and interfaces.With the recent development of biomaterials and biodevices with reduced dimensionality, it is critical to comprehend protein adhesion processes to nanoscale solid surfaces, especially those

  19. Clustering amino acid contents of protein domains: biochemical functions of proteins and implications for origin of biological macromolecules.

    PubMed

    Torshin, I Y

    2001-04-01

    Structural classes of protein domains correlate with their amino acid compositions. Several successful algorithms (that use only amino acid composition) have been elaborated for the prediction of structural class or potential biochemical significance. This work deals with dynamic classification (clustering) of the domains on the basis of their amino acid composition. Amino acid contents of domains from a non-redundant PDB set were clustered in 20-dimensional space of amino acid contents. Despite the variations of an empirical parameter and non-redundancy of the set, only one large cluster (tens-hundreds of proteins) surrounded by hundreds of small clusters (1-5 proteins), was identified. The core of the largest cluster contains at least 64% DNA (nucleotide)-interacting protein domains from various sources. About 90% of the proteins of the core are intracellular proteins. 83% of the DNA/nucleotide interacting domains in the core belong to the mixed alpha-beta folds (a+b, a/b), 14% are all-alpha (mostly helices) and all-beta (mostly beta-strands) proteins. At the same time, when core domains that belong to one organism (E.coli) are considered, over 80% of them prove to be DNA/nucleotide interacting proteins. The core is compact: amino acid contents of domains from the core lie in relatively narrow and specific ranges. The core also contains several Fe-S cluster-binding domains, amino acid contents of the core overlap with ferredoxin and CO-dehydrogenase clusters, the oldest known proteins. As Fe-S clusters are thought to be the first biocatalysts, the results are discussed in relation to contemporary experiments and models dealing with the origin of biological macromolecules. The origin of most primordial proteins is considered here to be a result of co-adsorption of nucleotides and amino acids on specific clays, followed by en-block polymerization of the adsorbed mixtures of amino acids.

  20. Atomic-Level Characterization of the Chain-Flipping Mechanism in Fatty-Acids Biosynthesis.

    PubMed

    Colizzi, Francesco; Masetti, Matteo; Recanatini, Maurizio; Cavalli, Andrea

    2016-08-01

    During fatty acids biosynthesis the elongating acyl chain is sequestered within the core of the highly conserved acyl carrier protein (ACP). At each catalytic step, the acyl intermediates are transiently delivered from ACP to the active site of the enzymatic counterparts and, at the same time, are protected from the solvent to prevent nonselective reactivity. Yet, the molecular determinants of such a universal transition-termed chain flipping-remain poorly understood. Here we capture the atomic-level details of the chain-flipping mechanism by using metadynamics simulations. We observe the fatty-acid chain gliding through the protein-protein interface with barely 30% of its surface exposed to water molecules. The small ACP's helix III acts as gatekeeper of the process, and we find its conformational plasticity critical for a successful substrate transfer. The results are in agreement with a wide range of experimental observations and provide unprecedented insight on the molecular determinants and driving forces of the chain-flipping process. PMID:27409360

  1. Atomic-Level Characterization of the Chain-Flipping Mechanism in Fatty-Acids Biosynthesis.

    PubMed

    Colizzi, Francesco; Masetti, Matteo; Recanatini, Maurizio; Cavalli, Andrea

    2016-08-01

    During fatty acids biosynthesis the elongating acyl chain is sequestered within the core of the highly conserved acyl carrier protein (ACP). At each catalytic step, the acyl intermediates are transiently delivered from ACP to the active site of the enzymatic counterparts and, at the same time, are protected from the solvent to prevent nonselective reactivity. Yet, the molecular determinants of such a universal transition-termed chain flipping-remain poorly understood. Here we capture the atomic-level details of the chain-flipping mechanism by using metadynamics simulations. We observe the fatty-acid chain gliding through the protein-protein interface with barely 30% of its surface exposed to water molecules. The small ACP's helix III acts as gatekeeper of the process, and we find its conformational plasticity critical for a successful substrate transfer. The results are in agreement with a wide range of experimental observations and provide unprecedented insight on the molecular determinants and driving forces of the chain-flipping process.

  2. Lower serum uric acid level predicts mortality in dialysis patients

    PubMed Central

    Bae, Eunjin; Cho, Hyun-Jeong; Shin, Nara; Kim, Sun Moon; Yang, Seung Hee; Kim, Dong Ki; Kim, Yong-Lim; Kang, Shin-Wook; Yang, Chul Woo; Kim, Nam Ho; Kim, Yon Su; Lee, Hajeong

    2016-01-01

    Abstract We evaluated the impact of serum uric acid (SUA) on mortality in patients with chronic dialysis. A total of 4132 adult patients on dialysis were enrolled prospectively between August 2008 and September 2014. Among them, we included 1738 patients who maintained dialysis for at least 3 months and had available SUA in the database. We categorized the time averaged-SUA (TA-SUA) into 5 groups: <5.5, 5.5–6.4, 6.5–7.4, 7.5–8.4, and ≥8.5 mg/dL. Cox regression analysis was used to calculate the hazard ratio (HR) of all-cause mortality according to SUA group. The mean TA-SUA level was slightly higher in men than in women. Patients with lower TA-SUA level tended to have lower body mass index (BMI), phosphorus, serum albumin level, higher proportion of diabetes mellitus (DM), and higher proportion of malnourishment on the subjective global assessment (SGA). During a median follow-up of 43.9 months, 206 patients died. Patients with the highest SUA had a similar risk to the middle 3 TA-SUA groups, but the lowest TA-SUA group had a significantly elevated HR for mortality. The lowest TA-SUA group was significantly associated with increased all-cause mortality (adjusted HR, 1.720; 95% confidence interval, 1.007–2.937; P = 0.047) even after adjusting for demographic, comorbid, nutritional covariables, and medication use that could affect SUA levels. This association was prominent in patients with well nourishment on the SGA, a preserved serum albumin level, a higher BMI, and concomitant DM although these parameters had no significant interaction in the TA-SUA-mortality relationship except DM. In conclusion, a lower TA-SUA level <5.5 mg/dL predicted all-cause mortality in patients with chronic dialysis. PMID:27310949

  3. Amino Acid Residues in the ω-Minus Region Participate in Cellular Localization of Yeast Glycosylphosphatidylinositol-Attached Proteins

    PubMed Central

    Hamada, Kenji; Terashima, Hiromichi; Arisawa, Mikio; Yabuki, Nami; Kitada, Kunio

    1999-01-01

    The final destination of glycosylphosphatidylinositol (GPI)-attached proteins in Saccharomyces cerevisiae is the plasma membrane or the cell wall. Two kinds of signals have been proposed for their cellular localization: (i) the specific amino acid residues V, I, or L at the site 4 or 5 amino acids upstream of the GPI attachment site (the ω site) and Y or N at the site 2 amino acids upstream of the ω site for cell wall localization and (ii) dibasic residues in the region upstream of the ω site (the ω-minus region) for plasma membrane localization. The relationships between these amino acid residues and efficiencies of cell wall incorporation were examined by constructing fusion reporter proteins from open reading frames encoding putative GPI-attached proteins. The levels of incorporation were high in the constructs containing the specific amino acid residues and quite low in those containing two basic amino acid residues in the ω-minus region. With constructs that contained neither specific residues nor two basic residues, levels of incorporation were moderate. These correlations clearly suggest that GPI-attached proteins have two different signals which act positively or negatively in cell wall incorporation for their cellular localization. PMID:10383953

  4. Omega-3 fatty acid deficiency disrupts endocytosis, neuritogenesis, and mitochondrial protein pathways in the mouse hippocampus

    PubMed Central

    English, Jane A.; Harauma, Akiko; Föcking, Melanie; Wynne, Kieran; Scaife, Caitriona; Cagney, Gerard; Moriguchi, Toru; Cotter, David R.

    2013-01-01

    Omega-3 fatty acid (n-3 FA) deficiency is an environmental risk factor for schizophrenia, yet characterization of the consequences of deficiency at the protein level in the brain is limited. We aimed to identify the protein pathways disrupted as a consequence of chronic n-3 deficiency in the hippocampus of mice. Fatty acid analysis of the hippocampus following chronic dietary deficiency revealed a 3-fold decrease (p < 0.001) in n-3 FA levels. Label free LC-MS/MS analysis identified and profiled 1008 proteins, of which 114 were observed to be differentially expressed between n-3 deficient and control groups (n = 8 per group). The cellular processes that were most implicated were neuritogenesis, endocytosis, and exocytosis, while specific protein pathways that were most significantly dysregulated were mitochondrial dysfunction and clathrin mediated endocytosis (CME). In order to characterize whether these processes and pathways are ones influenced by antipsychotic medication, we used LC-MS/MS to test the differential expression of these 114 proteins in the hippocampus of mice chronically treated with the antipsychotic agent haloperidol. We observed 23 of the 114 proteins to be differentially expressed, 17 of which were altered in the opposite direction to that observed following n-3 deficiency. Overall, our findings point to disturbed synaptic function, neuritogenesis, and mitochondrial function as a consequence of dietary deficiency in n-3 FA. This study greatly aids our understanding of the molecular mechanism by which n-3 deficiency impairs normal brain function, and provides clues as to how n-3 FA exert their therapeutic effect in early psychosis. PMID:24194745

  5. Higher transcription levels in ascorbic acid biosynthetic and recycling genes were associated with higher ascorbic acid accumulation in blueberry.

    PubMed

    Liu, Fenghong; Wang, Lei; Gu, Liang; Zhao, Wei; Su, Hongyan; Cheng, Xianhao

    2015-12-01

    In our preliminary study, the ripe fruits of two highbush blueberry (Vaccinium corymbosum L.) cultivars, cv 'Berkeley' and cv 'Bluecrop', were found to contain different levels of ascorbic acid. However, factors responsible for these differences are still unknown. In the present study, ascorbic acid content in fruits was compared with expression profiles of ascorbic acid biosynthetic and recycling genes between 'Bluecrop' and 'Berkeley' cultivars. The results indicated that the l-galactose pathway was the predominant route of ascorbic acid biosynthesis in blueberry fruits. Moreover, higher expression levels of the ascorbic acid biosynthetic genes GME, GGP, and GLDH, as well as the recycling genes MDHAR and DHAR, were associated with higher ascorbic acid content in 'Bluecrop' compared with 'Berkeley', which indicated that a higher efficiency ascorbic acid biosynthesis and regeneration was likely to be responsible for the higher ascorbic acid accumulation in 'Bluecrop'.

  6. Chemical cross-linking/mass spectrometry targeting acidic residues in proteins and protein complexes.

    PubMed

    Leitner, Alexander; Joachimiak, Lukasz A; Unverdorben, Pia; Walzthoeni, Thomas; Frydman, Judith; Förster, Friedrich; Aebersold, Ruedi

    2014-07-01

    The study of proteins and protein complexes using chemical cross-linking followed by the MS identification of the cross-linked peptides has found increasingly widespread use in recent years. Thus far, such analyses have used almost exclusively homobifunctional, amine-reactive cross-linking reagents. Here we report the development and application of an orthogonal cross-linking chemistry specific for carboxyl groups. Chemical cross-linking of acidic residues is achieved using homobifunctional dihydrazides as cross-linking reagents and a coupling chemistry at neutral pH that is compatible with the structural integrity of most protein complexes. In addition to cross-links formed through insertion of the dihydrazides with different spacer lengths, zero-length cross-link products are also obtained, thereby providing additional structural information. We demonstrate the application of the reaction and the MS identification of the resulting cross-linked peptides for the chaperonin TRiC/CCT and the 26S proteasome. The results indicate that the targeting of acidic residues for cross-linking provides distance restraints that are complementary and orthogonal to those obtained from lysine cross-linking, thereby expanding the yield of structural information that can be obtained from cross-linking studies and used in hybrid modeling approaches. PMID:24938783

  7. Chemical cross-linking/mass spectrometry targeting acidic residues in proteins and protein complexes

    PubMed Central

    Leitner, Alexander; Joachimiak, Lukasz A.; Unverdorben, Pia; Walzthoeni, Thomas; Frydman, Judith; Förster, Friedrich; Aebersold, Ruedi

    2014-01-01

    The study of proteins and protein complexes using chemical cross-linking followed by the MS identification of the cross-linked peptides has found increasingly widespread use in recent years. Thus far, such analyses have used almost exclusively homobifunctional, amine-reactive cross-linking reagents. Here we report the development and application of an orthogonal cross-linking chemistry specific for carboxyl groups. Chemical cross-linking of acidic residues is achieved using homobifunctional dihydrazides as cross-linking reagents and a coupling chemistry at neutral pH that is compatible with the structural integrity of most protein complexes. In addition to cross-links formed through insertion of the dihydrazides with different spacer lengths, zero-length cross-link products are also obtained, thereby providing additional structural information. We demonstrate the application of the reaction and the MS identification of the resulting cross-linked peptides for the chaperonin TRiC/CCT and the 26S proteasome. The results indicate that the targeting of acidic residues for cross-linking provides distance restraints that are complementary and orthogonal to those obtained from lysine cross-linking, thereby expanding the yield of structural information that can be obtained from cross-linking studies and used in hybrid modeling approaches. PMID:24938783

  8. Characterization of protein expression levels with label-free detected reverse phase protein arrays.

    PubMed

    Guo, Xuexue; Deng, Yihong; Zhu, Chenggang; Cai, Junlong; Zhu, Xiangdong; Landry, James P; Zheng, Fengyun; Cheng, Xunjia; Fei, Yiyan

    2016-09-15

    In reverse-phase protein arrays (RPPA), one immobilizes complex samples (e.g., cellular lysate, tissue lysate or serum etc.) on solid supports and performs parallel reactions of antibodies with immobilized protein targets from the complex samples. In this work, we describe a label-free detection of RPPA that enables quantification of RPPA data and thus facilitates comparison of studies performed on different samples and on different solid supports. We applied this detection platform to characterization of phosphoserine aminotransferase (PSAT) expression levels in Acanthamoeba lysates treated with artemether and the results were confirmed by Western blot studies. PMID:27372609

  9. Small acid soluble proteins for rapid spore identification.

    SciTech Connect

    Branda, Steven S.; Lane, Todd W.; VanderNoot, Victoria A.; Jokerst, Amanda S.

    2006-12-01

    This one year LDRD addressed the problem of rapid characterization of bacterial spores such as those from the genus Bacillus, the group that contains pathogenic spores such as B. anthracis. In this effort we addressed the feasibility of using a proteomics based approach to spore characterization using a subset of conserved spore proteins known as the small acid soluble proteins or SASPs. We proposed developing techniques that built on our previous expertise in microseparations to rapidly characterize or identify spores. An alternative SASP extraction method was developed that was amenable to both the subsequent fluorescent labeling required for laser-induced fluorescence detection and the low ionic strength requirements for isoelectric focusing. For the microseparations, both capillary isoelectric focusing and chip gel electrophoresis were employed. A variety of methods were evaluated to improve the molecular weight resolution for the SASPs, which are in a molecular weight range that is not well resolved by the current methods. Isoelectric focusing was optimized and employed to resolve the SASPs using UV absorbance detection. Proteomic signatures of native wild type Bacillus spores and clones genetically engineered to produce altered SASP patterns were assessed by slab gel electrophoresis, capillary isoelectric focusing with absorbance detection as well as microchip based gel electrophoresis employing sensitive laser-induced fluorescence detection.

  10. Coffee bean arabinogalactans: acidic polymers covalently linked to protein.

    PubMed

    Redgwell, Robert J; Curti, Delphine; Fischer, Monica; Nicolas, Pierre; Fay, Laurent B

    2002-02-11

    The arabinogalactan content of green coffee beans (Coffea arabica var. Yellow Caturra) was released by a combination of chemical extraction and enzymatic hydrolysis of the mannan-cellulose component of the wall. Several arabinogalactan fractions were isolated, purified by gel-permeation and ion-exchange chromatography and characterised by compositional and linkage analysis. The AG fractions contained between 6 and 8% glucuronic acid, and gave a positive test for the beta-glucosyl-Yariv reagent, a stain specific for arabinogalactan-proteins. The protein component accounted for between 0.5 and 2.0% of the AGPs and contained between 7 and 12% hydroxyproline. The AG moieties displayed considerable heterogeneity with regard to their degree of arabinosylation and the extent and composition of their side-chains. They possessed a MW average of 650 kDa which ranged between 150 and 2000 kDa. An investigation of the structural features of the major AG fraction, released following enzymatic hydrolysis of the mannan-cellulose polymers, allowed a partial structure of coffee arabinogalactan to be proposed.

  11. Chitosan grafted low molecular weight polylactic acid for protein encapsulation and burst effect reduction.

    PubMed

    Di Martino, Antonio; Kucharczyk, Pavel; Zednik, Jiri; Sedlarik, Vladimir

    2015-12-30

    Chitosan and chitosan-grafted polylactic acid as a matrix for BSA encapsulation in a nanoparticle structure were prepared through a polyelectrolyte complexation method with dextran sulfate. Polylactic acid was synthetized via a polycondensation reaction using the non-metal-based initiator methanesulfonic acid and grafted to the chitosan backbone by a coupling reaction, with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide as the condensing agent. The effect of concentration of the polymer matrix utilized herein on particle diameter, ζ-potential, encapsulation efficiency, and the release kinetic of the model protein bovine serum albumin at differing pH levels was investigated. The influence of pH and ionic strength on the behavior of the nanoparticles prepared was also researched. Results showed that grafting polylactic acid to chitosan chains reduced the initial burst effect in the kinetics of BSA release from the structure of the nanoparticles. Furthermore, a rise in encapsulation efficiency of the bovine serum albumin and diminishment in nanoparticle diameter were observed due to chitosan modification. The results suggest that both polymers actually show appreciable encapsulation efficiency; and release rate of BSA. CS-g-PLA is more suitable than unmodified CS as a carrier for controlled protein delivery. PMID:26453778

  12. Nutritional consequences of the reactions between proteins and oxidized polyphenolic acids.

    PubMed

    Hurrell, R F; Finot, P A

    1984-01-01

    The chemical and enzymatic browning reactions of plant polyphenols and their effects on amino acids and proteins are reviewed. A model system of casein and oxidizing caffeic acid has been studied in more detail. The effects of pH, time, caffeic acid level and the presence or not of tyrosinase on the decrease of FDNB-reactive lysine are described. The chemical loss of lysine, methionine and tryptophan and the change in the bioavailability of these amino acids to rats has been evaluated in two systems: pH 7.0 with tyrosinase and pH 10.0 without tyrosinase. At pH 10.0, reactive lysine was more reduced. At pH 7.0 plus tyrosinase methionine was more extensively oxidized to its sulphoxide. Tryptophan was not chemically reduced under either condition. At pH 10.0 there was a decrease in the protein digestibility which was responsible for a corresponding reduction in tryptophan availability and partly responsible for lower methionine availability. Metabolic transit of casein labelled with tritiated lysine treated under the same conditions indicated that the lower lysine availability in rats was due to a lower digestibility of the lysine-caffeoquinone complexes. PMID:6496220

  13. Chitosan grafted low molecular weight polylactic acid for protein encapsulation and burst effect reduction.

    PubMed

    Di Martino, Antonio; Kucharczyk, Pavel; Zednik, Jiri; Sedlarik, Vladimir

    2015-12-30

    Chitosan and chitosan-grafted polylactic acid as a matrix for BSA encapsulation in a nanoparticle structure were prepared through a polyelectrolyte complexation method with dextran sulfate. Polylactic acid was synthetized via a polycondensation reaction using the non-metal-based initiator methanesulfonic acid and grafted to the chitosan backbone by a coupling reaction, with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide as the condensing agent. The effect of concentration of the polymer matrix utilized herein on particle diameter, ζ-potential, encapsulation efficiency, and the release kinetic of the model protein bovine serum albumin at differing pH levels was investigated. The influence of pH and ionic strength on the behavior of the nanoparticles prepared was also researched. Results showed that grafting polylactic acid to chitosan chains reduced the initial burst effect in the kinetics of BSA release from the structure of the nanoparticles. Furthermore, a rise in encapsulation efficiency of the bovine serum albumin and diminishment in nanoparticle diameter were observed due to chitosan modification. The results suggest that both polymers actually show appreciable encapsulation efficiency; and release rate of BSA. CS-g-PLA is more suitable than unmodified CS as a carrier for controlled protein delivery.

  14. Prolonged fasting identifies heat shock protein 10 as a Sirtuin 3 substrate: elucidating a new mechanism linking mitochondrial protein acetylation to fatty acid oxidation enzyme folding and function.

    PubMed

    Lu, Zhongping; Chen, Yong; Aponte, Angel M; Battaglia, Valentina; Gucek, Marjan; Sack, Michael N

    2015-01-23

    Although Sirtuin 3 (SIRT3), a mitochondrially enriched deacetylase and activator of fat oxidation, is down-regulated in response to high fat feeding, the rate of fatty acid oxidation and mitochondrial protein acetylation are invariably enhanced in this dietary milieu. These paradoxical data implicate that additional acetylation modification-dependent levels of regulation may be operational under nutrient excess conditions. Because the heat shock protein (Hsp) Hsp10-Hsp60 chaperone complex mediates folding of the fatty acid oxidation enzyme medium-chain acyl-CoA dehydrogenase, we tested whether acetylation-dependent mitochondrial protein folding contributes to this regulatory discrepancy. We demonstrate that Hsp10 is a functional SIRT3 substrate and that, in response to prolonged fasting, SIRT3 levels modulate mitochondrial protein folding. Acetyl mutagenesis of Hsp10 lysine 56 alters Hsp10-Hsp60 binding, conformation, and protein folding. Consistent with Hsp10-Hsp60 regulation of fatty acid oxidation enzyme integrity, medium-chain acyl-CoA dehydrogenase activity and fat oxidation are elevated by Hsp10 acetylation. These data identify acetyl modification of Hsp10 as a nutrient-sensing regulatory node controlling mitochondrial protein folding and metabolic function. PMID:25505263

  15. Source of amino acids for tRNA acylation. Implications for measurement of protein synthesis.

    PubMed Central

    Barnes, D M; Calvert, C C; Klasing, K C

    1992-01-01

    Estimates of protein-synthesis rates using radioisotopes require accurate measurement of the specific radioactivity of the label in protein and in the precursor pool over time. Although the extracellular and intracellular pools of amino acids are easiest to sample, the tRNA pool is the direct precursor and is the appropriate pool for sampling. To test if the intracellular or extracellular pools reflect the tRNA specific radioactivity, a chicken macrophage cell line was incubated in medium containing either 0.23 mM-leucine and 14.5 microCi of [3H]leucine (tracer dose) or 2.3 microM-leucine plus 145.0 microCi of [3H]leucine (flooding dose). At both leucine levels, the tRNA specific radioactivity reached a plateau quickly, but did not equilibrate with either the extracellular or intracellular specific radioactivity within 30 min, and remained closer to that of protein. In a second experiment, proteins in chicken macrophages were labelled with [3H]leucine for 2 days. Labelling medium was removed, and the cells were washed free of residual free [3H]leucine and incubated with medium containing either 0.23 mM- or 2.3 mM-leucine (unlabelled). The specific radioactivity of leucyl-tRNA leucine reached a plateau within 2 min and remained considerably closer to that in the protein than that in intracellular or extracellular pools for at least 60 min. These results suggest that amino acids from protein degradation are a primary source for charging tRNA. When protein-synthesis rates are estimated by label incorporation, use of extracellular or intracellular specific-radioactivity values result in a marked underestimation. PMID:1575701

  16. Use of stable isotopes to assess protein and amino acid metabolism in children and adolescents: a brief review.

    PubMed

    Darmaun, Dominique; Mauras, Nelly

    2005-01-01

    As protein accretion is a prerequisite for growth, studying the mechanisms by which nutrients and hormones promote protein gain is of the utmost relevance to paediatric endocrinology. Tracers are ideally suited for the assessment of protein and amino acid kinetics in vivo, as they provide an estimate of synthesis and turnover. Current tracer approaches in children and adolescents utilize stable isotopes, 'heavier' forms of elements that have one or several extra neutrons in the nucleus. Such isotopes are already present at low, but significant, levels in all tissues and foodstuffs, are not radioactive and are devoid of any known side-effects when present in small amounts. L-[1-(13)C] labelled leucine, given as a 4- to 6-h intravenous infusion, has become the method of choice to assess whole-body protein kinetics. After infusion, any 13C-leucine that is oxidized appears in the breath as 13CO2, whereas the remainder is incorporated into body proteins through protein synthesis. The isotope enrichments are determined by isotope ratio mass spectrometry and gas chromatography mass spectrometry, and absolute rates of whole-body protein synthesis, oxidation, and breakdown can be extrapolated. This approach has been used extensively to investigate the regulation of protein kinetics by nutrients and by hormones. Attempts have also been made to measure amino acid/protein metabolism in selected body compartments, and to measure the kinetics of specific tissue proteins, for example, muscle, gut, or plasma proteins.

  17. Expression of Pokeweed Antiviral Protein in Transgenic Plants Induces Virus Resistance in Grafted Wild-Type Plants Independently of Salicylic Acid Accumulation and Pathogenesis-Related Protein Synthesis.

    PubMed Central

    Smirnov, S.; Shulaev, V.; Tumer, N. E.

    1997-01-01

    Pokeweed antiviral protein (PAP), a 29-kD protein isolated from Phytolacca americana, inhibits translation by catalytically removing a specific adenine residue from the large rRNA of the 60S subunit of eukaryotic ribosomes. Transgenic tobacco (Nicotiana tabacum) plants expressing PAP or a variant (PAP-v) were shown to be resistant to a broad spectrum of plant viruses. Expression of PAP-v in transgenic plants induces synthesis of pathogenesis-related proteins and a very weak (<2-fold) increase in salicylic acid levels. Using reciprocal grafting experiments, we demonstrate here that transgenic tobacco rootstocks expressing PAP-v induce resistance to tobacco mosaic virus infection in both N. tabacum NN and nn scions. Increased resistance to potato virus X was also observed in N. tabacum nn scions grafted on transgenic rootstocks. PAP expression was not detected in the wild-type scions or rootstocks that showed virus resistance, nor was there any increase in salicylic acid levels or pathogenesis-related protein synthesis. Grafting experiments with transgenic plants expressing an inactive PAP mutant demonstrated that an intact active site of PAP is necessary for induction of virus resistance in wild-type scions. These results indicate that enzymatic activity of PAP is responsible for generating a signal that renders wild-type scions resistant to virus infection in the absence of increased salicylic acid levels and pathogenesis-related protein synthesis. PMID:12223762

  18. C-reactive protein levels in hereditary angioedema

    PubMed Central

    Hofman, Z L M; Relan, A; Hack, C E

    2014-01-01

    Hereditary angioedema (HAE) patients experience recurrent episodes of angioedema attacks that can be painful, disfiguring and even life-threatening. The disorder results from a mutation in the gene that controls the synthesis of C1-inhibitor (C1INH). C1INH is a major regulator of activation of the contact system. It is often assumed that attacks results from uncontrolled local activation of the contact system with subsequent formation of bradykinin. To evaluate the involvement of inflammatory reactions in HAE, we analysed C-reactive protein (CRP) levels. HAE patients included in a clinical database of recombinant human C1-inhibitor (rhC1INH) studies were evaluated. For the current study we analysed CRP levels when patients were asymptomatic, during a clinical attack and in a follow-up period, and correlated these with the clinical manifestations of the attack. Data from 68 HAE patients were analysed and included CRP levels on 273 occasions. While asymptomatic, 20% of the patients analysed had increased CRP. At the onset of the attack (P = 0·049) and during the next 24 h CRP rose significantly (P = 0·002) in patients with an abdominal location, and post-attack levels were significantly higher in these patients than in patients with attacks at other locations (P = 0·034). In conclusion, CRP levels are elevated in a substantial proportion of asymptomatic HAE patients. Levels of CRP increase significantly during an abdominal attack. These data suggest low-grade systemic inflammatory reactions in HAE patients as well as a triggering event for attacks that starts prior to symptom onset. PMID:24588117

  19. C-reactive protein levels in hereditary angioedema.

    PubMed

    Hofman, Z L M; Relan, A; Hack, C E

    2014-07-01

    Hereditary angioedema (HAE) patients experience recurrent episodes of angioedema attacks that can be painful, disfiguring and even life-threatening. The disorder results from a mutation in the gene that controls the synthesis of C1-inhibitor (C1INH). C1INH is a major regulator of activation of the contact system. It is often assumed that attacks results from uncontrolled local activation of the contact system with subsequent formation of bradykinin. To evaluate the involvement of inflammatory reactions in HAE, we analysed C-reactive protein (CRP) levels. HAE patients included in a clinical database of recombinant human C1-inhibitor (rhC1INH) studies were evaluated. For the current study we analysed CRP levels when patients were asymptomatic, during a clinical attack and in a follow-up period, and correlated these with the clinical manifestations of the attack. Data from 68 HAE patients were analysed and included CRP levels on 273 occasions. While asymptomatic, 20% of the patients analysed had increased CRP. At the onset of the attack (P = 0·049) and during the next 24 h CRP rose significantly (P = 0·002) in patients with an abdominal location, and post-attack levels were significantly higher in these patients than in patients with attacks at other locations (P = 0·034). In conclusion, CRP levels are elevated in a substantial proportion of asymptomatic HAE patients. Levels of CRP increase significantly during an abdominal attack. These data suggest low-grade systemic inflammatory reactions in HAE patients as well as a triggering event for attacks that starts prior to symptom onset.

  20. Metabolic labeling with stable isotope nitrogen (15N) to follow amino acid and protein turnover of three plastid proteins in Chlamydomonas reinhardtii

    PubMed Central

    2014-01-01

    Background The length of time that a protein remains available to perform its function is significantly influenced by its turnover rate. Knowing the turnover rate of proteins involved in different processes is important to determining how long a function might progress even when the stimulus has been removed and no further synthesis of the particular proteins occurs. In this article, we describe the use of 15N-metabolic labeling coupled to GC-MS to follow the turnover of free amino acids and LC-MS/MS to identify and LC-MS to follow the turnover of specific proteins in Chlamydomonas reinhardtii. Results To achieve the metabolic labeling, the growth medium was formulated with standard Tris acetate phosphate medium (TAP) in which14NH4Cl was replaced with 15NH415NO3 and (14NH4)6Mo7O24.4H2O was replaced with Na2MoO4.2H2O. This medium designated 15N-TAP allowed CC-125 algal cells to grow normally. Mass isotopic distribution revealed successful 15N incorporation into 13 amino acids with approximately 98% labeling efficiency. Tryptic digestion of the 55 kDa SDS-PAGE bands from 14N- and 15N-labeled crude algal protein extracts followed by LC-MS/MS resulted in the identification of 27 proteins. Of these, five displayed peptide sequence confidence levels greater than 95% and protein sequence coverage greater than 25%. These proteins were the RuBisCo large subunit, ATP synthase CF1 alpha and beta subunits, the mitochondrial protein (F1F0 ATP synthase) and the cytosolic protein (S-adenosyl homocysteine hydroxylase). These proteins were present in both labeled and unlabeled samples. Once the newly synthesized 15N-labeled free amino acids and proteins obtained maximum incorporation of the 15N-label, turnover rates were determined after transfer of cells into 14N-TAP medium. The t½ values were determined for the three plastid proteins (RuBisCo, ATP synthase CF1 alpha and beta) by following the reduction of the 15N-fractional abundance over time. Conclusion We describe a more

  1. Regulation of laminin and entactin mRNA levels by retinoic acid and dibutyryl cyclic AMP

    SciTech Connect

    Durkin, M.E.; Phillips, S.L.; Carlin, B.E.; Merlie, J.P.; Chung, A.E.

    1986-05-01

    Retinoic acid and dibutyryl cAMP induced F9 embryonal carcinoma cells to differentiate to parietal endoderm; the morphological changes were accompanied by the increased synthesis of the basement membrane glycoproteins laminin and entactin. cDNA clones have been isolated for the A (400 kD), B1 (220 kD), and B2 (205 kD) chains of laminin. Northern blot analysis indicated that the A, B1, and B2 chains were encoded by RNA species of 9.8, 6.0, and 8.0 kb, respectively. The kinetics of induction of the laminin mRNAs were studied by dot-blotting dilutions of RNA extracted from F9 cells cultured in retinoic acid and dibutyryl cAMP for increasing amounts of time and hybridizing to /sup 32/P-labeled recombinant plasmids. Very low levels of the A and B chain RNAs were found in uninduced cells, and a large increase occurred between 48 and 72 hr of growth in retinoic acid and dibutyryl cAMP. A cDNA clone was also obtained for entactin, a 150 kD glycoprotein that forms a complex with laminin. Retinoic acid and dibutyryl cAMP treatment also increased the amount of entactin RNA in F9 cells. These results suggested that a common mechanism may exist for the coordinate regulation of the 4 basement membrane protein genes during differentiation.

  2. Phenylketonuria: protein content and amino acids profile of dishes for phenylketonuric patients. The relevance of phenylalanine.

    PubMed

    Pimentel, Filipa B; Alves, Rita C; Costa, Anabela S G; Torres, Duarte; Almeida, Manuela F; Oliveira, M Beatriz P P

    2014-04-15

    Phenylketonuria is an inborn error of metabolism, involving, in most cases, a deficient activity of phenylalanine hydroxylase. Neonatal diagnosis and a prompt special diet (low phenylalanine and natural-protein restricted diets) are essential to the treatment. The lack of data concerning phenylalanine contents of processed foodstuffs is an additional limitation for an already very restrictive diet. Our goals were to quantify protein (Kjeldahl method) and amino acid (18) content (HPLC/fluorescence) in 16 dishes specifically conceived for phenylketonuric patients, and compare the most relevant results with those of several international food composition databases. As might be expected, all the meals contained low protein levels (0.67-3.15 g/100 g) with the highest ones occurring in boiled rice and potatoes. These foods also contained the highest amounts of phenylalanine (158.51 and 62.65 mg/100 g, respectively). In contrast to the other amino acids, it was possible to predict phenylalanine content based on protein alone. Slight deviations were observed when comparing results with the different food composition databases.

  3. Chenodeoxycholic Acid Reduces Hypoxia Inducible Factor-1α Protein and Its Target Genes.

    PubMed

    Moon, Yunwon; Choi, Su Mi; Chang, Soojeong; Park, Bongju; Lee, Seongyeol; Lee, Mi-Ock; Choi, Hueng-Sik; Park, Hyunsung

    2015-01-01

    This study evaluated HIF-1α inhibitors under different hypoxic conditions, physiological hypoxia (5% O2) and severe hypoxia (0.1% O2). We found that chenodeoxy cholic acid (CDCA) reduced the amount of HIF-1α protein only under physiological hypoxia but not under severe hypoxia without decreasing its mRNA level. By using a proteasome inhibitor MG132 and a translation inhibitor cyclohexamide, we showed that CDCA reduced HIF-1α protein by decreasing its translation but not by enhancing its degradation. The following findings indicated that farnesoid X receptor (FXR), a CDCA receptor and its target gene, Small heterodimer partner (SHP) are not involved in this effect of CDCA. Distinctly from CDCA, MG132 prevented SHP and an exogenous FXR agonist, GW4064 from reducing HIF-1α protein. Furthermore a FXR antagonist, guggulsterone failed to prevent CDCA from decreasing HIF-1α protein. Furthermore, guggulsterone by itself reduced HIF-1α protein even in the presence of MG132. These findings suggested that CDCA and guggulsterone reduced the translation of HIF-1α in a mechanism which FXR and SHP are not involved. This study reveals novel therapeutic functions of traditional nontoxic drugs, CDCA and guggulsterone, as inhibitors of HIF-1α protein.

  4. Chenodeoxycholic Acid Reduces Hypoxia Inducible Factor-1α Protein and Its Target Genes

    PubMed Central

    Moon, Yunwon; Choi, Su Mi; Chang, Soojeong; Park, Bongju; Lee, Seongyeol; Lee, Mi-Ock; Choi, Hueng-Sik; Park, Hyunsung

    2015-01-01

    This study evaluated HIF-1α inhibitors under different hypoxic conditions, physiological hypoxia (5% O2) and severe hypoxia (0.1% O2). We found that chenodeoxy cholic acid (CDCA) reduced the amount of HIF-1α protein only under physiological hypoxia but not under severe hypoxia without decreasing its mRNA level. By using a proteasome inhibitor MG132 and a translation inhibitor cyclohexamide, we showed that CDCA reduced HIF-1α protein by decreasing its translation but not by enhancing its degradation. The following findings indicated that farnesoid X receptor (FXR), a CDCA receptor and its target gene, Small heterodimer partner (SHP) are not involved in this effect of CDCA. Distinctly from CDCA, MG132 prevented SHP and an exogenous FXR agonist, GW4064 from reducing HIF-1α protein. Furthermore a FXR antagonist, guggulsterone failed to prevent CDCA from decreasing HIF-1α protein. Furthermore, guggulsterone by itself reduced HIF-1α protein even in the presence of MG132. These findings suggested that CDCA and guggulsterone reduced the translation of HIF-1α in a mechanism which FXR and SHP are not involved. This study reveals novel therapeutic functions of traditional nontoxic drugs, CDCA and guggulsterone, as inhibitors of HIF-1α protein. PMID:26098428

  5. Heavy metal levels and delta-amino-levulinic acid dehydrase levels in peripheral polyneuropathy.

    PubMed

    Melgaard, B; Clausen, J; Rastogi, S C

    1976-04-01

    On the basis of assay of heavy metals in whole blood (lead, cadimum, chromium, copper, nickel and manganese) and delta-amino-levulinic-acid dehydrase (ALA-D) and ALA in urine in a normal Danish population, the levels of these clinco-chemical factors were assayed in 23 patients with peripheral neuropathy of unknown etiology. All patients studied showed electro-physiological sign of denervation and/or reduced motor or sensoric nerve conduction velocity. Cadmium and manganese were never found to be increased. In all but four patients, an increase of one or more heavy metals was found. Ten patients showed raised levels of two or more metals, the dominant metal being lead (10 cases), nine patients showed increased in chromium. A significant corrleation was found between increasing lead levels and decreasing ALA-D activity. Although normal concentrations of manganese were found, correlation analysis revealed a significant correlation between increased manganese and decreased ALA-D. The raised values of heavy metals could not be traced to occupational or other exposure to heavy metals and the increased values were not related to tobacco consumption. The findings are discussed in relation to known data on neuropathy and the results seem to indicate a multifactorial patholgenesis of the disease. Among factors contributing to the precipitation of the syndrome may be raised levels of heavy metals.

  6. Biology Teacher and Expert Opinions about Computer Assisted Biology Instruction Materials: A Software Entitled Nucleic Acids and Protein Synthesis

    ERIC Educational Resources Information Center

    Hasenekoglu, Ismet; Timucin, Melih

    2007-01-01

    The aim of this study is to collect and evaluate opinions of CAI experts and biology teachers about a high school level Computer Assisted Biology Instruction Material presenting computer-made modelling and simulations. It is a case study. A material covering "Nucleic Acids and Protein Synthesis" topic was developed as the "case". The goal of the…

  7. Experimental Evolution of a Green Fluorescent Protein Composed of 19 Unique Amino Acids without Tryptophan

    NASA Astrophysics Data System (ADS)

    Kawahara-Kobayashi, Akio; Hitotsuyanagi, Mitsuhiro; Amikura, Kazuaki; Kiga, Daisuke

    2014-04-01

    At some stage of evolution, genes of organisms may have encoded proteins that were synthesized using fewer than 20 unique amino acids. Similar to evolution of the natural 19-amino-acid proteins GroEL/ES, proteins composed of 19 unique amino acids would have been able to evolve by accumulating beneficial mutations within the 19-amino-acid repertoire encoded in an ancestral genetic code. Because Trp is thought to be the last amino acid included in the canonical 20-amino-acid repertoire, this late stage of protein evolution could be mimicked by experimental evolution of 19-amino-acid proteins without tryptophan (Trp). To further understand the evolution of proteins, we tried to mimic the evolution of a 19-amino-acid protein involving the accumulation of beneficial mutations using directed evolution by random mutagenesis on the whole targeted gene sequence. We created active 19-amino-acid green fluorescent proteins (GFPs) without Trp from a poorly fluorescent 19-amino-acid mutant, S1-W57F, by using directed evolution with two rounds of mutagenesis and selection. The N105I and S205T mutations showed beneficial effects on the S1-W57F mutant. When these two mutations were combined on S1-W57F, we observed an additive effect on the fluorescence intensity. In contrast, these mutations showed no clear improvement individually or in combination on GFPS1, which is the parental GFP mutant composed of 20 amino acids. Our results provide an additional example for the experimental evolution of 19-amino-acid proteins without Trp, and would help understand the mechanisms underlying the evolution of 19-amino-acid proteins. (236 words)

  8. [Relationship between the included levels of coffee pulp and the protein content in rations for monogastric animals].

    PubMed

    Gómez-Brenes, R A; Bendaña, G; González, J M; Braham, J E; Bressani, R

    1985-09-01

    The purpose of this research was to determine the effect of including fresh and ensilaged coffee pulp in rations for monogastric animals, and find the best protein and coffee pulp levels in rations for rats. Fresh coffee pulp and pulp ensilaged for 12 months were used; both kinds of pulp were sun-dried before incorporating them into the rations. The chemical analyses of the pulps revealed a lower content in caffeine, tannins, chlorogenic acid and caffeic acid in the ensilaged pulp than in fresh coffee pulp. Thirty-two experimental rations were prepared, 16 with fresh coffee pulp and 16 with the ensilaged by-product, distributed into four different protein levels (10, 15, 20 and 25%), and three levels of pulp (15, 30 and 45%) for each protein level. The rations thus prepared were fed to Wistar albino rats for a six-week period. The parameters used to measure the effect of the two types of pulp were mortality rate, food consumption, weight gain, food conversion and apparent digestibility of the rations. Ensilaged pulp had a higher nutritive value, lower toxicity and better digestibility than fresh pulp. The increase in the protein level of the ration resulted in partial protection against the negative effects of coffee pulp on the performance of animals, since this improved as the protein level of the ration increased.

  9. Effects of folic acid supplementation on serum homocysteine and lipoprotein (a) levels during pregnancy

    PubMed Central

    Hekmati Azar Mehrabani, Zohreh; Ghorbanihaghjo, Amir; Sayyah Melli, Manizheh; Hamzeh-Mivehroud, Maryam; Fathi Maroufi, Nazila; Bargahi, Nasrin; Bannazadeh Amirkhiz, Maryam; Rashtchizadeh, Nadereh

    2015-01-01

    Introduction:There are many ideas concerning the etiology and pathogenesis of preeclampsia including endothelial dysfunction, inflammation and angiogenesis. Elevated levels of total homocysteine (Hcy) and lipoprotein (a) [Lp(a)] are risk factors for endothelial dysfunction. This study aimed to evaluate the effect of high dose folic acid (FA) on serum Hcy and Lp(a) concentrations with respect to methylenetetrahydrofolate reductase (MTHFR) polymorphisms 677C→T during pregnancy. Methods: In a prospective uncontrolled intervention, 90 pregnant women received 5 mg FA supplementation before pregnancy till 36th week of pregnancy. The MTHFR polymorphisms 677C→T, serum lactate dehydrogenase activity, urine protein and creatinine concentrations were measured before starting folic acid administration. Serum levels of Hcy and Lp(a) were determined before and after completion of folic acid supplementation period. Results: Supplementation of the patients with FA for 36 week decreased the median (minimum– maximum) levels of serum Hcy from 11.40 μmol/L (4.40-28.70) to 9.70 (1.60-20.80) μmol/L (p=0.001). There was no significant change in serum Lp(a) after FA supplementation (p=0.17). The overall prevalence of genotypes in pregnant women that were under study for MTHFR C677T polymorphism was 53.3% CC, 26.7% CT and 20.0% TT. There was no correlation between decreasing level of serum Hcy in the patients receiving FA and MTHFR polymorphisms. Conclusion:Although FA supplementation decreased serum levels of Hcy in different MTHFR genotypes, serum Lp(a) was not changed by FA supplements. Our data suggests that FA supplementation effects on serum Hcy is MTHFR genotype independent in pregnant women. PMID:26929921

  10. Identification of Acidic pH-dependent Ligands of Pentameric C-reactive Protein*

    PubMed Central

    Hammond, David J.; Singh, Sanjay K.; Thompson, James A.; Beeler, Bradley W.; Rusiñol, Antonio E.; Pangburn, Michael K.; Potempa, Lawrence A.; Agrawal, Alok

    2010-01-01

    C-reactive protein (CRP) is a phylogenetically conserved protein; in humans, it is present in the plasma and at sites of inflammation. At physiological pH, native pentameric CRP exhibits calcium-dependent binding specificity for phosphocholine. In this study, we determined the binding specificities of CRP at acidic pH, a characteristic of inflammatory sites. We investigated the binding of fluid-phase CRP to six immobilized proteins: complement factor H, oxidized low-density lipoprotein, complement C3b, IgG, amyloid β, and BSA immobilized on microtiter plates. At pH 7.0, CRP did not bind to any of these proteins, but, at pH ranging from 5.2 to 4.6, CRP bound to all six proteins. Acidic pH did not monomerize CRP but modified the pentameric structure, as determined by gel filtration, 1-anilinonaphthalene-8-sulfonic acid-binding fluorescence, and phosphocholine-binding assays. Some modifications in CRP were reversible at pH 7.0, for example, the phosphocholine-binding activity of CRP, which was reduced at acidic pH, was restored after pH neutralization. For efficient binding of acidic pH-treated CRP to immobilized proteins, it was necessary that the immobilized proteins, except factor H, were also exposed to acidic pH. Because immobilization of proteins on microtiter plates and exposure of immobilized proteins to acidic pH alter the conformation of immobilized proteins, our findings suggest that conformationally altered proteins form a CRP-ligand in acidic environment, regardless of the identity of the protein. This ligand binding specificity of CRP in its acidic pH-induced pentameric state has implications for toxic conditions involving protein misfolding in acidic environments and favors the conservation of CRP throughout evolution. PMID:20843812

  11. The second amino acid of alfalfa mosaic virus coat protein is critical for coat protein-mediated protection.

    PubMed Central

    Tumer, N E; Kaniewski, W; Haley, L; Gehrke, L; Lodge, J K; Sanders, P

    1991-01-01

    Transgenic plants expressing the coat protein (CP) of alfalfa mosaic virus (AIMV) are resistant to infection by AIMV. A mutation was introduced into the second amino acid of the cDNA for the CP of AIMV. Three different transgenic tobacco lines expressing the mutant CP and two different transgenic tobacco lines expressing the wild-type CP at similar levels were challenged with AIMV virions and viral RNA. Whereas the lines expressing the wild-type CP were highly resistant to infection by AIMV virions and viral RNA, the lines expressing the mutant CP were susceptible to infection by both. The binding affinity of the mutant and the wild-type CPs for the 3' terminal protein binding site on AIMV RNAs was similar, as determined by electrophoretic mobility shift assay. A mixture of AIMV genomic RNAs 1-3 was infectious on the plants expressing the mutant CP but not on vector control plants or plants expressing the wild-type CP, indicating that the mutant CP can activate the AIMV genomic RNAs for infection. These results demonstrate that the second amino acid of the AIMV CP is critical for protection from AIMV but not for the initial interaction between the AIMV RNA and CP, suggesting that this initial interaction does not play a major role in CP-mediated protection. Images PMID:11607167

  12. Prebiotic Synthesis of Hydrophobic and Protein Amino Acids

    PubMed Central

    Ring, David; Wolman, Yecheskel; Friedmann, Nadav; Miller, Stanley L.

    1972-01-01

    The formation of amino acids by the action of electric discharges on a mixture of methane, nitrogen, and water with traces of ammonia was studied in detail. The presence of glycine, alanine, α-amino-n-butyric acid, α-aminoisobutyric acid, valine, norvaline, isovaline, leucine, isoleucine, alloisoleucine, norleucine, proline, aspartic acid, glutamic acid, serine, threonine, allothreonine, α-hydroxy-γ-aminobutyric acid, and α,γ-diaminobutyric acid was confirmed by ion-exchange chromatography and gas chromatography-mass spectrometry. All of the primary α-amino acids found in the Murchison Meteorite have been synthesized by this electric discharge experiment. PMID:4501592

  13. Microwave-assisted acid and base hydrolysis of intact proteins containing disulfide bonds for protein sequence analysis by mass spectrometry.

    PubMed

    Reiz, Bela; Li, Liang

    2010-09-01

    Controlled hydrolysis of proteins to generate peptide ladders combined with mass spectrometric analysis of the resultant peptides can be used for protein sequencing. In this paper, two methods of improving the microwave-assisted protein hydrolysis process are described to enable rapid sequencing of proteins containing disulfide bonds and increase sequence coverage, respectively. It was demonstrated that proteins containing disulfide bonds could be sequenced by MS analysis by first performing hydrolysis for less than 2 min, followed by 1 h of reduction to release the peptides originally linked by disulfide bonds. It was shown that a strong base could be used as a catalyst for microwave-assisted protein hydrolysis, producing complementary sequence information to that generated by microwave-assisted acid hydrolysis. However, using either acid or base hydrolysis, amide bond breakages in small regions of the polypeptide chains of the model proteins (e.g., cytochrome c and lysozyme) were not detected. Dynamic light scattering measurement of the proteins solubilized in an acid or base indicated that protein-protein interaction or aggregation was not the cause of the failure to hydrolyze certain amide bonds. It was speculated that there were some unknown local structures that might play a role in preventing an acid or base from reacting with the peptide bonds therein.

  14. Effect of branched chain amino acid infusions on body protein metabolism in cirrhosis of liver.

    PubMed Central

    Wright, P D; Holdsworth, J D; Dionigi, P; Clague, M B; James, O F

    1986-01-01

    Thirty seven patients with established cirrhosis of the liver were subjected to measurement of body protein metabolism using L-(1-14C) labelled leucine as a tracer. The effects of disease severity and those of solutions containing 0%, 16%, 35%, 53%, and 100% branched chain amino acids were evaluated. Significant increases in protein synthesis were noted with solutions containing 35%, 53%, and 100% branched chain amino acids, but in patients receiving 100% branched chain amino acids without additional essential amino acid supplement the increase in synthesis was matched by a significant increase in protein breakdown. Protein balance was thus improved only in patients receiving 35% and 53% branched chain amino acids. It was concluded that the high increase in protein breakdown in patients receiving 100% branched chain amino acids was undesirable, and such a solution should not be recommended for clinical use. PMID:3539714

  15. The action of exogenous gibberellic acid on protein and mRNA in germinating castor bean seeds.

    PubMed

    Martin, C; Northcote, D H

    1982-03-01

    Gibberellic acid (GA3) stimulates water uptake in castor beans and increases the activity of certain enzymes associated with lipid mobilisation.The effect of the GA3 on the enzymes is possibly due to a general effect of the growth substance on protein synthesis. Gibberellic acid advanced the appearance of rRNA and poly (A(+))RNA in castor bean endosperms without specifically stimulating the synthesis of particular mRNA species. Thus these increased levels of mRNA and rRNA may act synergistically to affect the rate of a predetermined pattern of protein synthesis.

  16. CMP‑N‑acetylneuraminic acid synthetase interacts with fragile X related protein 1.

    PubMed

    Ma, Yun; Tian, Shuai; Wang, Zongbao; Wang, Changbo; Chen, Xiaowei; Li, Wei; Yang, Yang; He, Shuya

    2016-08-01

    Fragile X mental retardation protein (FMRP), fragile X related 1 protein (FXR1P) and FXR2P are the members of the FMR protein family. These proteins contain two KH domains and a RGG box, which are characteristic of RNA binding proteins. The absence of FMRP, causes fragile X syndrome (FXS), the leading cause of hereditary mental retardation. FXR1P is expressed throughout the body and important for normal muscle development, and its absence causes cardiac abnormality. To investigate the functions of FXR1P, a screen was performed to identify FXR1P‑interacting proteins and determine the biological effect of the interaction. The current study identified CMP‑N‑acetylneuraminic acid synthetase (CMAS) as an interacting protein using the yeast two‑hybrid system, and the interaction between FXR1P and CMAS was validated in yeast using a β‑galactosidase assay and growth studies with selective media. Furthermore, co‑immunoprecipitation was used to analyze the FXR1P/CMAS association and immunofluorescence microscopy was performed to detect expression and intracellular localization of the proteins. The results of the current study indicated that FXR1P and CMAS interact, and colocalize in the cytoplasm and the nucleus of HEK293T and HeLa cells. Accordingly, a fragile X related 1 (FXR1) gene overexpression vector was constructed to investigate the effect of FXR1 overexpression on the level of monosialotetrahexosylganglioside 1 (GM1). The results of the current study suggested that FXR1P is a tissue‑specific regulator of GM1 levels in SH‑SY5Y cells, but not in HEK293T cells. Taken together, the results initially indicate that FXR1P interacts with CMAS, and that FXR1P may enhance the activation of sialic acid via interaction with CMAS, and increase GM1 levels to affect the development of the nervous system, thus providing evidence for further research into the pathogenesis of FXS.

  17. Liquid chromatographic analysis of phosphoamino acids at femtomole level using chemical derivatization with N-hydroxysuccinimidyl fluorescein-O-acetate.

    PubMed

    Deng, Ying-Hua; Li, Rong-Jun; Zhang, Hua-Shan; Du, Xiao-Lan; Wang, Hong

    2007-10-01

    Phosphorylation of amino acid residues in proteins plays a major role in biological systems. In this paper, a reversed-phase high performance liquid chromatographic (HPLC) method based on chemical derivatization has been described for the separation and quantification of phosphoamino acids at femtomole level, using fluorimetric detection (FLD). The protocol involved pre-column derivatization of phosphoamino acids with N-hydroxysuccinimidyl fluorescein-O-acetate (SIFA) and subsequent separation on ZORBAX Eclipse XDB-C8 column. Several experimental factors that influenced derivatization and separation were carefully investigated. The derivatization was performed at 40 degrees C for 40 min in borate buffer (pH 8.5). Under the optimum conditions, phosphoserine (P-Ser), phosphothreonine (P-Thr) and phosphotyrosine (P-Tyr) were satisfactorily separated in 8 min. The detection limits (signal-to-noise ratio=3) for the phosphoamino acids could reach 10-20 fmol, which was the lowest value reported for HPLC methods and comparable to those obtained by capillary electrophoresis with laser-induced fluorescence (CE-LIF) detection methods. The proposed method has been validated and used to characterize the phosphoamino acids in the hydrolyzed phosphorylated protein samples. The results clearly demonstrated the potential of this technique to study phosphoamino acids as well as provided a new analytical methodology that should be applicable to the study of phosphorylation of protein in biological system.

  18. Honey bee protein atlas at organ-level resolution.

    PubMed

    Chan, Queenie W T; Chan, Man Yi; Logan, Michelle; Fang, Yuan; Higo, Heather; Foster, Leonard J

    2013-11-01

    Genome sequencing has provided us with gene lists but cannot tell us where and how their encoded products work together to support life. Complex organisms rely on differential expression of subsets of genes/proteins in organs and tissues, and, in concert, evolved to their present state as they function together to improve an organism's overall reproductive fitness. Proteomics studies of individual organs help us understand their basic functions, but this reductionist approach misses the larger context of the whole organism. This problem could be circumvented if all the organs in an organism were comprehensively studied by the same methodology and analyzed together. Using honey bees (Apis mellifera L.) as a model system, we report here an initial whole proteome of a complex organism, measuring 29 different organ/tissue types among the three honey bee castes: queen, drone, and worker. The data reveal that, e.g., workers have a heightened capacity to deal with environmental toxins and queens have a far more robust pheromone detection system than their nestmates. The data also suggest that workers altruistically sacrifice not only their own reproductive capacity but also their immune potential in favor of their queen. Finally, organ-level resolution of protein expression offers a systematic insight into how organs may have developed.

  19. Honey bee protein atlas at organ-level resolution.

    PubMed

    Chan, Queenie W T; Chan, Man Yi; Logan, Michelle; Fang, Yuan; Higo, Heather; Foster, Leonard J

    2013-11-01

    Genome sequencing has provided us with gene lists but cannot tell us where and how their encoded products work together to support life. Complex organisms rely on differential expression of subsets of genes/proteins in organs and tissues, and, in concert, evolved to their present state as they function together to improve an organism's overall reproductive fitness. Proteomics studies of individual organs help us understand their basic functions, but this reductionist approach misses the larger context of the whole organism. This problem could be circumvented if all the organs in an organism were comprehensively studied by the same methodology and analyzed together. Using honey bees (Apis mellifera L.) as a model system, we report here an initial whole proteome of a complex organism, measuring 29 different organ/tissue types among the three honey bee castes: queen, drone, and worker. The data reveal that, e.g., workers have a heightened capacity to deal with environmental toxins and queens have a far more robust pheromone detection system than their nestmates. The data also suggest that workers altruistically sacrifice not only their own reproductive capacity but also their immune potential in favor of their queen. Finally, organ-level resolution of protein expression offers a systematic insight into how organs may have developed. PMID:23878156

  20. Cullin-3 protein expression levels correlate with breast cancer progression

    PubMed Central

    Haagenson, Kelly K.; Tait, Larry; Wang, Juan; Shekhar, Malathy P.; Polin, Lisa; Chen, Wei; Wu, Gen Sheng

    2012-01-01

    Cullin-3 is a component of the Cullin-Ring ubiquitin ligase (CRL) family that plays an important role in mediating protein degradation. Deregulation of Cullin-3 expression has been observed in human cancers; however, a role for Cullin-3 in tumor progression has not been previously recognized. Using the MCF10DCIS.com human breast cancer xenograft model, we show that Cullin-3 is increasingly expressed during progression from comedo ductal carcinoma in situ (DCIS) to invasive carcinomas. Cullin-3 protein is not detected in early lesions but is noticeably increased in DCIS tumors and significantly overexpressed in invasive cancers. In experimental metastasis assays, high expression of Cullin-3 was observed in the lung site. Importantly, Cullin-3 staining is detected in human breast cancer tissues, not in normal breast tissues and its expression level positively correlates with tumor stage. These data suggest that Cullin-3 may play an important role in tumor progression from DCIS to invasive cancer and may serve as a biomarker for the diagnosis of aggressive breast cancer. PMID:22825334

  1. Synthesis of alanyl nucleobase amino acids and their incorporation into proteins.

    PubMed

    Talukder, Poulami; Dedkova, Larisa M; Ellington, Andrew D; Yakovchuk, Petro; Lim, Jaebum; Anslyn, Eric V; Hecht, Sidney M

    2016-09-15

    Proteins which bind to nucleic acids and regulate their structure and functions are numerous and exceptionally important. Such proteins employ a variety of strategies for recognition of the relevant structural elements in their nucleic acid substrates, some of which have been shown to involve rather subtle interactions which might have been difficult to design from first principles. In the present study, we have explored the preparation of proteins containing unnatural amino acids having nucleobase side chains. In principle, the introduction of multiple nucleobase amino acids into the nucleic acid binding domain of a protein should enable these modified proteins to interact with their nucleic acid substrates using Watson-Crick and other base pairing interactions. We describe the synthesis of five alanyl nucleobase amino acids protected in a fashion which enabled their attachment to a suppressor tRNA, and their incorporation into each of two proteins with acceptable efficiencies. The nucleobases studied included cytosine, uracil, thymine, adenine and guanine, i.e. the major nucleobase constituents of DNA and RNA. Dihydrofolate reductase was chosen as one model protein to enable direct comparison of the facility of incorporation of the nucleobase amino acids with numerous other unnatural amino acids studied previously. The Klenow fragment of DNA polymerase I was chosen as a representative DNA binding protein whose mode of action has been studied in detail. PMID:27452282

  2. Synthesis of alanyl nucleobase amino acids and their incorporation into proteins.

    PubMed

    Talukder, Poulami; Dedkova, Larisa M; Ellington, Andrew D; Yakovchuk, Petro; Lim, Jaebum; Anslyn, Eric V; Hecht, Sidney M

    2016-09-15

    Proteins which bind to nucleic acids and regulate their structure and functions are numerous and exceptionally important. Such proteins employ a variety of strategies for recognition of the relevant structural elements in their nucleic acid substrates, some of which have been shown to involve rather subtle interactions which might have been difficult to design from first principles. In the present study, we have explored the preparation of proteins containing unnatural amino acids having nucleobase side chains. In principle, the introduction of multiple nucleobase amino acids into the nucleic acid binding domain of a protein should enable these modified proteins to interact with their nucleic acid substrates using Watson-Crick and other base pairing interactions. We describe the synthesis of five alanyl nucleobase amino acids protected in a fashion which enabled their attachment to a suppressor tRNA, and their incorporation into each of two proteins with acceptable efficiencies. The nucleobases studied included cytosine, uracil, thymine, adenine and guanine, i.e. the major nucleobase constituents of DNA and RNA. Dihydrofolate reductase was chosen as one model protein to enable direct comparison of the facility of incorporation of the nucleobase amino acids with numerous other unnatural amino acids studied previously. The Klenow fragment of DNA polymerase I was chosen as a representative DNA binding protein whose mode of action has been studied in detail.

  3. Chicoric Acid Levels in Basil (Ocimum basilicum L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recently, we reported the presence of chicoric acid in basil leaves (confirmed by co-chromatography with purchased standard). Chicoric acid being the chief phenolic of the Echinacea purpurea plant which is popularly consumed as a dietary supplement. For this study, basil products commonly purchased ...

  4. Cell-cycle-dependent changes in ceramide levels preceding retinoblastoma protein dephosphorylation in G2/M.

    PubMed Central

    Lee, J Y; Leonhardt, L G; Obeid, L M

    1998-01-01

    Ceramide functions as a growth-inhibitory lipid-signalling molecule and might have a role in mediating the effects of extracellular agents on cell growth, differentiation and senescence. Here we investigate the roles of ceramide in cell cycle progression. With the use of the model of serum withdrawal, we were able to synchronize Wi-38 human diploid fibroblasts at different stages of cell cycle. Serum stimulation resulted in G0 to G1/S progression as determined by flow cytometric analysis and [3H]thymidine incorporation. Analyses of endogenous ceramide levels demonstrated that ceramide levels remained relatively constant on serum stimulation, indicating that ceramide might not be critical during G1/S transition. Treating exponentially growing Wi-38 human diploid fibroblasts with nocodazole led to cell cycle arrest at the G2/M phase of the cell cycle; 2 h after the removal of nocodazole, retinoblastoma (Rb) protein became dephosphorylated and the cells exited from G2/M and moved to the G1 phase of the new cycle. When cells were released from G2/M block by nocodazole, and before Rb protein dephosphorylation, endogenous ceramide levels transiently increased up to 2-fold at 0.5 h after the removal of nocodazole. Fumonisin B1, an inhibitor of ceramide synthase, inhibited the elevation of ceramide levels. Desipramine and SR33557, both acid sphingomyelinase inhibitors, did not have an appreciable effect on the elevation of ceramide levels. Furthermore, fumonisin B1 inhibited Rb protein dephosphorylation induced by endogenous ceramide but not by exogenous ceramide. These results demonstrate for the first time changes in ceramide during cell cycle progression and suggest that ceramide synthesized de novo might function as an endogenous modulator of Rb protein and cell cycle progression. PMID:9716505

  5. Non-protein amino acids in Australian acacia seed: implications for food security and recommended processing methods to reduce djenkolic acid.

    PubMed

    Boughton, Berin A; Reddy, Priyanka; Boland, Martin P; Roessner, Ute; Yates, Peter

    2015-07-15

    Seed of Australian acacia species, Acacia colei, Acacia elecantha, Acacia torulosa, Acacia turmida and Acacia saligna, were analysed for the presence of toxic non-protein amino acids and the levels of essential amino acids. Amines were derivatised with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate before analysis using liquid chromatography electrospray ionisation triple quadrupole mass spectrometry (LC-ESI-QQQ-MS). Multiple reaction monitoring (MRM) with optimised transitions and collision energies for each analyte were employed. The known nephrotoxic compound djenkolic acid was found to be present at elevated levels in all species tested. The lowest levels were in A. colei (0.49% w/w) and the highest in A. saligna (1.85% w/w). Observed levels of djenkolic acid are comparable to measured and reported levels found in the djenkol bean. Subsequent testing of seed processing methods showed djenkolic acid levels can be significantly reduced by over 90% by dry roasting at 180 °C rendering the seed safe for human consumption.

  6. Studies of composition and major protein level in milk and colostrum of mares.

    PubMed

    Pecka, Ewa; Dobrzański, Zbigniew; Zachwieja, Andrzej; Szulc, Tadeusz; Czyż, Katarzyna

    2012-02-01

    The aim of the study was to determine the changes in composition and physicochemical features (pH, density, thermostability and acidity) of mare colostrum and milk, and of protein fraction contribution (serum albumin, β-casein, γ-casein, α-lactalbumin, G class immunoglobulins) depending on lactation stage. The research material was colostrum and milk samples from 12 Arabian mares. Colostrum samples were collected within 2 h after parturition and milk samples were collected twice, in the 3rd and 6th weeks of lactation. The level of basic milk components decreased significantly (only lactose content increased) as compared to colostrum. Total bacteria count and somatic cell count decreased significantly with an increase in resistance and urea level. The changes observed were connected to differentiated contribution of particular protein fractions and their relative proportions. Lower levels of γ-casein (P ≤ 0.05), β-casein, serum albumin as well as α-lactalbumin were observed in colostrum as compared to those in milk. Any relationship between lactation stage and β-casein content was observed. Serum albumin and α-lactalbumin content increased in subsequent milkings. The level of G class immunoglobulins decreased significantly and its highest level was noted in colostrum. Any significant differences between the 3rd and 6th lactation weeks were obtained. PMID:22339698

  7. Copper uptake is required for pyrrolidine dithiocarbamate-mediated oxidation and protein level increase of p53 in cells.

    PubMed Central

    Furuta, Saori; Ortiz, Fausto; Zhu Sun, Xiu; Wu, Hsiao-Huei; Mason, Andrew; Momand, Jamil

    2002-01-01

    The p53 tumour-suppressor protein is a transcription factor that activates the expression of genes involved in cell cycle arrest, apoptosis and DNA repair. The p53 protein is vulnerable to oxidation at cysteine thiol groups. The metal-chelating dithiocarbamates, pyrrolidine dithiocarbamate (PDTC), diethyldithiocarbamate, ethylene(bis)dithiocarbamate and H(2)O(2) were tested for their oxidative effects on p53 in cultured human breast cancer cells. Only PDTC oxidized p53, although all oxidants tested increased the p53 level. Inductively coupled plasma MS analysis indicated that the addition of 60 microM PDTC increased the cellular copper concentration by 4-fold, which was the highest level of copper accumulated amongst all the oxidants tested. Bathocuproinedisulphonic acid, a membrane-impermeable Cu(I) chelator inhibited the PDTC-mediated copper accumulation. Bathocuproinedisulphonic acid as well as the hydroxyl radical scavenger d-mannitol inhibited the PDTC-dependent increase in p53 protein and oxidation. Our results show that a low level of copper accumulation in the range of 25-40 microg/g of cellular protein increases the steady-state levels of p53. At copper accumulation levels higher than 60 microg/g of cellular protein, p53 is oxidized. These results suggest that p53 is vulnerable to free radical-mediated oxidation at cysteine residues. PMID:11964141

  8. Sex Steroid Modulation of Fatty Acid Utilization and Fatty Acid Binding Protein Concentration in Rat Liver

    PubMed Central

    Ockner, Robert K.; Lysenko, Nina; Manning, Joan A.; Monroe, Scott E.; Burnett, David A.

    1980-01-01

    The mechanism by which sex steroids influence very low density hepatic lipoprotein triglyceride production has not been fully elucidated. In previous studies we showed that [14C]oleate utilization and incorporation into triglycerides were greater in hepatocyte suspensions from adult female rats than from males. The sex differences were not related to activities of the enzymes of triglyceride biosynthesis, whereas fatty acid binding protein (FABP) concentration in liver cytosol was greater in females. These findings suggested that sex differences in lipoprotein could reflect a sex steroid influence on the availability of fatty acids for hepatocellular triglyceride biosynthesis. In the present studies, sex steroid effects on hepatocyte [14C]oleate utilization and FABP concentration were investigated directly. Hepatocytes from immature (30-d-old) rats exhibited no sex differences in [14C]oleate utilization. With maturation, total [14C]oleate utilization and triglyceride biosynthesis increased moderately in female cells and decreased markedly in male cells; the profound sex differences in adults were maximal by age 60 d. Fatty acid oxidation was little affected. Rats were castrated at age 30 d, and received estradiol, testosterone, or no hormone until age 60 d, when hepatocyte [14C]oleate utilization was studied. Castration virtually eliminated maturational changes and blunted the sex differences in adults. Estradiol or testosterone largely reproduced the appropriate adult pattern of [14C]oleate utilization regardless of the genotypic sex of the treated animal. In immature females and males, total cytosolic FABP concentrations were similar. In 60-d-old animals, there was a striking correlation among all groups (females, males, castrates, and hormone-treated) between mean cytosolic FABP concentration on the one hand, and mean total [14C]oleate utilization (r = 0.91) and incorporation into triglycerides (r = 0.94) on the other. In 30-d-old animals rates of [14C

  9. High levels of gene expression explain the strong evolutionary constraint of mitochondrial protein-coding genes.

    PubMed

    Nabholz, Benoit; Ellegren, Hans; Wolf, Jochen B W

    2013-02-01

    The nearly neutral theory of molecular evolution has been widely accepted as the guiding principle for understanding how selection affects gene sequence evolution. One of its central predictions is that the rate at which proteins evolve should negatively scale with effective population size (N(e)). In contrast to the expectation of reduced selective constraint in the mitochondrial genome following from its lower N(e), we observe what can be interpreted as the opposite: for a taxonomically diverse set of organisms (birds, mammals, insects, and nematodes), mitochondrially encoded protein-coding genes from the oxidative phosphorylation pathway (mtOXPHOS; n = 12-13) show markedly stronger signatures of purifying selection (illustrated by low d(N)/d(S)) than their nuclear counterparts interacting in the same pathway (nuOXPHOS; n: ∼75). To understand these unexpected evolutionary dynamics, we consider a number of structural and functional parameters including gene expression, hydrophobicity, transmembrane position, gene ontology, GC content, substitution rate, proportion of amino acids in transmembrane helices, and protein-protein interaction. Across all taxa, unexpectedly large differences in gene expression levels (RNA-seq) between nuclear and mitochondrially encoded genes, and to a lower extent hydrophobicity, explained most of the variation in d(N)/d(S). Similarly, differences in d(N)/d(S) between functional OXPHOS protein complexes could largely be explained by gene expression differences. Overall, by including gene expression and other functional parameters, the unexpected mitochondrial evolutionary dynamics can be understood. Our results not only reaffirm the link between gene expression and protein evolution but also open new questions about the functional role of expression level variation between mitochondrial genes. PMID:23071102

  10. High levels of gene expression explain the strong evolutionary constraint of mitochondrial protein-coding genes.

    PubMed

    Nabholz, Benoit; Ellegren, Hans; Wolf, Jochen B W

    2013-02-01

    The nearly neutral theory of molecular evolution has been widely accepted as the guiding principle for understanding how selection affects gene sequence evolution. One of its central predictions is that the rate at which proteins evolve should negatively scale with effective population size (N(e)). In contrast to the expectation of reduced selective constraint in the mitochondrial genome following from its lower N(e), we observe what can be interpreted as the opposite: for a taxonomically diverse set of organisms (birds, mammals, insects, and nematodes), mitochondrially encoded protein-coding genes from the oxidative phosphorylation pathway (mtOXPHOS; n = 12-13) show markedly stronger signatures of purifying selection (illustrated by low d(N)/d(S)) than their nuclear counterparts interacting in the same pathway (nuOXPHOS; n: ∼75). To understand these unexpected evolutionary dynamics, we consider a number of structural and functional parameters including gene expression, hydrophobicity, transmembrane position, gene ontology, GC content, substitution rate, proportion of amino acids in transmembrane helices, and protein-protein interaction. Across all taxa, unexpectedly large differences in gene expression levels (RNA-seq) between nuclear and mitochondrially encoded genes, and to a lower extent hydrophobicity, explained most of the variation in d(N)/d(S). Similarly, differences in d(N)/d(S) between functional OXPHOS protein complexes could largely be explained by gene expression differences. Overall, by including gene expression and other functional parameters, the unexpected mitochondrial evolutionary dynamics can be understood. Our results not only reaffirm the link between gene expression and protein evolution but also open new questions about the functional role of expression level variation between mitochondrial genes.

  11. Spaceflight effects on adult rat muscle protein, nucleic acids, and amino acids

    NASA Technical Reports Server (NTRS)

    Steffen, J. M.; Musacchia, X. J.

    1986-01-01

    Exposure to conditions of weightlessness has been associated with decrements in muscle mass and strength. The present studies were undertaken to determine muscle responses at the cellular level. Male Sprague-Dawley rats (360-410 g) were exposed to 7 days of weightlessness during the Spacelab-3 shuttle flight (May 1985). Animals were killed 12 h postflight, and soleus (S), gastrocnemius (G), and extensor digitorum longus (EDL) muscles were excised. Muscle protein, RNA, and DNA were extracted and quantified. Differential muscle atrophy was accompanied by a significant (P less than 0.05) reduction in total protein only in S muscles. There were no significant changes in protein concentration (mg/g) in the muscles examined. In S muscles from flight animals, sarcoplasmic protein accounted for a significantly greater proportion of total protein that in ground controls (37.5 vs. 32.5%). Tissue concentrations (nmol/g) of asparagine-aspartate, glutamine-glutamate, glycine, histidine, and lysine were significantly reduced (from 17 to 63%) in S muscles from flight animals, but only glutamine-glutamate levels were decreased in the G and EDL. Muscle DNA content (microgram) was unchanged in the tissues examined, but S muscle DNA concentration (micrograms/mg) increased 27%. RNA content (micrograms) was significantly (P less than 0.025) reduced in S (-28%) and G(-22%) muscles following spaceflight. These results identify specific alterations in rat skeletal muscle during short term (7-day) exposure to weightlessness and compare favorably with observations previously obtained from ground-based suspension simulations.

  12. The Citrus transcription factor, CitERF13, regulates citric acid accumulation via a protein-protein interaction with the vacuolar proton pump, CitVHA-c4

    PubMed Central

    Li, Shao-jia; Yin, Xue-ren; Xie, Xiu-lan; Allan, Andrew C.; Ge, Hang; Shen, Shu-ling; Chen, Kun-song

    2016-01-01

    Organic acids are essential to fruit flavor. The vacuolar H+ transporting adenosine triphosphatase (V-ATPase) plays an important role in organic acid transport and accumulation. However, less is known of V-ATPase interacting proteins and their relationship with organic acid accumulation. The relationship between V-ATPase and citric acid was investigated, using the citrus tangerine varieties ‘Ordinary Ponkan (OPK)’ and an early maturing mutant ‘Zaoshu Ponkan (ZPK)’. Five V-ATPase genes (CitVHA) were predicted as important to citric acid accumulation. Among the genes, CitVHA-c4 was observed, using a yeast two-hybrid screen, to interact at the protein level with an ethylene response factor, CitERF13. This was verified using bimolecular fluorescence complementation assays. A similar interaction was also observed between Arabidopsis AtERF017 (a CitERF13 homolog) and AtVHA-c4 (a CitVHA-c4 homolog). A synergistic effect on citric acid levels was observed between V-ATPase proteins and interacting ERFs when analyzed using transient over-expression in tobacco and Arabidopsis mutants. Furthermore, the transcript abundance of CitERF13 was concomitant with CitVHA-c4. CitERF13 or AtERF017 over-expression leads to significant citric acid accumulation. This accumulation was abolished in an AtVHA-c4 mutant background. ERF-VHA interactions appear to be involved in citric acid accumulation, which was observed in both citrus and Arabidopsis. PMID:26837571

  13. Probing Protein Channel Dynamics At The Single Molecule Level.

    NASA Astrophysics Data System (ADS)

    Lee, M. Ann; Dunn, Robert C.

    1997-03-01

    It would be difficult to overstate the importance played by protein ion channels in cellular function. These macromolecular pores allow the passage of ions across the cellular membrane and play indispensable roles in all aspects of neurophysiology. While the patch-clamp technique continues to provide elegant descriptions of the kinetic processes involved in ion channel gating, the associated conformational changes remain a mystery. We are using the spectroscopic capabilities and single molecule fluorescence sensitivity of near-field scanning optical microscopy (NSOM) to probe these dynamics at the single channel level. Using a newly developed cantilevered NSOM probe capable of probing soft biological samples with single molecule fluorescence sensitivity, we have begun mapping the location of single NMDA receptors in intact rat cortical neurons with <100 nm spatial resolution. We will also present recent results exploring the conformational changes accompanying activation of nuclear pore channels located in the nuclear membrane of Xenopus oocytes. Our recent NSOM and AFM measurements on single nuclear pore complexes reveal large conformational changes taking place upon activation, providing rich, new molecular level details of channel function.

  14. Effect of supplemental folic acid on valproic acid-induced embryotoxicity and tissue zinc levels in vivo.

    PubMed

    Hansen, D K; Grafton, T F; Dial, S L; Gehring, T A; Siitonen, P H

    1995-11-01

    Valproic acid (VPA) is an anti-convulsant drug known to cause spina bifida in humans. Administration of the vitamin, folic acid, has been shown to decrease the recurrence and possibly also the occurrence of neural tube defects, primarily spina bifida, in humans. Additionally, treatment with a derivative (folinic acid) of folic acid has been reported to decrease the frequency of VPA-induced exencephaly in mice treated with the drug in vivo. A protective effect by folinic acid has not been observed in vitro. The purpose of this investigation was to reexamine the ability of folinic acid to decrease the incidence of VPA-induced neural tube defects in vivo. We also examined the effect of increased intake of folic acid on zinc levels in various maternal and embryonic tissues. Folinic acid, whether administered by intraperitoneal injection or in osmotic mini-pumps, did not decrease the number of mouse fetuses with VPA-induced exencephaly. Dietary supplementation with 10-20 times the daily required intake of folic acid in rodents also failed to decrease the embryotoxicity of VPA. Such dietary supplementation had no effect on zinc levels in maternal liver, brain, or kidney, nor in embryonic tissues. These results indicate that folic acid is not able to reverse the embryotoxicity induced by the anticonvulsant, that there is no apparent effect of high dietary folate intake on maternal or embryonic zinc levels and suggest that folate is probably not involved in the mechanism of VPA-induced embryotoxicity. PMID:8838251

  15. Protein acetylation affects acetate metabolism, motility and acid stress response in Escherichia coli

    PubMed Central

    Castaño-Cerezo, Sara; Bernal, Vicente; Post, Harm; Fuhrer, Tobias; Cappadona, Salvatore; Sánchez-Díaz, Nerea C; Sauer, Uwe; Heck, Albert JR; Altelaar, AF Maarten; Cánovas, Manuel

    2014-01-01

    Although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. To interrogate its functionality, we analyzed the acetylome in Escherichia coli knockout mutants of cobB, the only known sirtuin-like deacetylase, and patZ, the best-known protein acetyltransferase. For four growth conditions, more than 2,000 unique acetylated peptides, belonging to 809 proteins, were identified and differentially quantified. Nearly 65% of these proteins are related to metabolism. The global activity of CobB contributes to the deacetylation of a large number of substrates and has a major impact on physiology. Apart from the regulation of acetyl-CoA synthetase, we found that CobB-controlled acetylation of isocitrate lyase contributes to the fine-tuning of the glyoxylate shunt. Acetylation of the transcription factor RcsB prevents DNA binding, activating flagella biosynthesis and motility, and increases acid stress susceptibility. Surprisingly, deletion of patZ increased acetylation in acetate cultures, which suggests that it regulates the levels of acetylating agents. The results presented offer new insights into functional roles of protein acetylation in metabolic fitness and global cell regulation. PMID:25518064

  16. Retinal functional alterations in mice lacking intermediate filament proteins glial fibrillary acidic protein and vimentin.

    PubMed

    Wunderlich, Kirsten A; Tanimoto, Naoyuki; Grosche, Antje; Zrenner, Eberhart; Pekny, Milos; Reichenbach, Andreas; Seeliger, Mathias W; Pannicke, Thomas; Perez, Maria-Thereza

    2015-12-01

    Vimentin (Vim) and glial fibrillary acidic protein (GFAP) are important components of the intermediate filament (IF) (or nanofilament) system of astroglial cells. We conducted full-field electroretinogram (ERG) recordings and found that whereas photoreceptor responses (a-wave) were normal in uninjured GFAP(-/-)Vim(-/-) mice, b-wave amplitudes were increased. Moreover, we found that Kir (inward rectifier K(+)) channel protein expression was reduced in the retinas of GFAP(-/-)Vim(-/-) mice and that Kir-mediated current amplitudes were lower in Müller glial cells isolated from these mice. Studies have shown that the IF system, in addition, is involved in the retinal response to injury and that attenuated Müller cell reactivity and reduced photoreceptor cell loss are observed in IF-deficient mice after experimental retinal detachment. We investigated whether the lack of IF proteins would affect cell survival in a retinal ischemia-reperfusion model. We found that although cell loss was induced in both genotypes, the number of surviving cells in the inner retina was lower in IF-deficient mice. Our findings thus show that the inability to produce GFAP and Vim affects normal retinal physiology and that the effect of IF deficiency on retinal cell survival differs, depending on the underlying pathologic condition.

  17. Potential Role for Purple Acid Phosphatase in the Dephosphorylation of Wall Proteins in Tobacco Cells1[W

    PubMed Central

    Kaida, Rumi; Serada, Satoshi; Norioka, Naoko; Norioka, Shigemi; Neumetzler, Lutz; Pauly, Markus; Sampedro, Javier; Zarra, Ignacio; Hayashi, Takahisa; Kaneko, Takako S.

    2010-01-01

    It is not yet known whether dephosphorylation of proteins catalyzed by phosphatases occurs in the apoplastic space. In this study, we found that tobacco (Nicotiana tabacum) purple acid phosphatase could dephosphorylate the phosphoryl residues of three apoplastic proteins, two of which were identified as α-xylosidase and β-glucosidase. The dephosphorylation and phosphorylation of recombinant α-xylosidase resulted in a decrease and an increase in its activity, respectively, when xyloglucan heptasaccharide was used as a substrate. Attempted overexpression of the tobacco purple acid phosphatase NtPAP12 in tobacco cells not only decreased the activity levels of the glycosidases but also increased levels of xyloglucan oligosaccharides and cello-oligosaccharides in the apoplast during the exponential phase. We suggest that purple acid phosphatase controls the activity of α-xylosidase and β-glucosidase, which are responsible for the degradation of xyloglucan oligosaccharides and cello-oligosaccharides in the cell walls. PMID:20357138

  18. Amino acid repletion does not decrease muscle protein catabolism during hemodialysis.

    PubMed

    Raj, Dominic S C; Adeniyi, Oladipo; Dominic, Elizabeth A; Boivin, Michel A; McClelland, Sandra; Tzamaloukas, Antonios H; Morgan, Nancy; Gonzales, Lawrence; Wolfe, Robert; Ferrando, Arny

    2007-06-01

    Intradialytic protein catabolism is attributed to loss of amino acids in the dialysate. We investigated the effect of amino acid infusion during hemodialysis (HD) on muscle protein turnover and amino acid transport kinetics by using stable isotopes of phenylalanine, leucine, and lysine in eight patients with end-stage renal disease (ESRD). Subjects were studied at baseline (pre-HD), 2 h of HD without amino acid infusion (HD-O), and 2 h of HD with amino acid infusion (HD+AA). Amino acid depletion during HD-O augmented the outward transport of amino acids from muscle into the vein. Increased delivery of amino acids to the leg during HD+AA facilitated the transport of amino acids from the artery into the intracellular compartment. Increase in muscle protein breakdown was more than the increase in synthesis during HD-O (46.7 vs. 22.3%, P < 0.001). Net balance (nmol.min(-1).100 ml (-1)) was more negative during HD-O compared with pre-HD (-33.7 +/- 1.5 vs. -6.0 +/- 2.3, P < 0.001). Despite an abundant supply of amino acids, the net balance (-16.9 +/- 1.8) did not switch from net release to net uptake. HD+AA induced a proportional increase in muscle protein synthesis and catabolism. Branched chain amino acid catabolism increased significantly from baseline during HD-O and did not decrease during HD+AA. Protein synthesis efficiency, the fraction of amino acid in the intracellular pool that is utilized for muscle protein synthesis decreased from 42.1% pre-HD to 33.7 and 32.6% during HD-O and HD+AA, respectively (P < 0.01). Thus amino acid repletion during HD increased muscle protein synthesis but did not decrease muscle protein breakdown. PMID:17264222

  19. Experimentally Testing the Hypothesis of a Limited Amino Acid Repertoire in Primitive Proteins

    NASA Astrophysics Data System (ADS)

    Akanuma, S.; Nakajima, Y.; Yokobori, S.; Yamagishi, A.

    2013-11-01

    It has been argued that a fewer amino acids were used in primitive proteins and later the repertoire increased up to 20. To test this hypothesis experimentally, we restricted the amino acid usage of a reconstructed, ancestral protein to reduced sets.

  20. Labeling proteins via hole burning of their aromatic amino acids: pressure tuning spectroscopy of BPTI.

    PubMed Central

    Stübner, Markus; Hecht, Christoph; Friedrich, Josef

    2002-01-01

    We demonstrate hole burning on a protein by using an intrinsic aromatic amino acid as a probe. The protein is bovine pancreatic trypsin inhibitor (BPTI), the labeled amino acid is tyrosine. Only one of the four tyrosines could be burned. As an application we present pressure tuning experiments from which the local compressibility around the burned tyrosine probe is determined. PMID:12496122

  1. Unravelling Protein-DNA Interactions at Molecular Level: A DFT and NCI Study.

    PubMed

    González, J; Baños, I; León, I; Contreras-García, J; Cocinero, E J; Lesarri, A; Fernández, J A; Millán, J

    2016-02-01

    Histone-DNA interactions were probed computationally at a molecular level, by characterizing the bimolecular clusters constituted by selected amino acid derivatives with polar (asparagine and glutamine), nonpolar (alanine, valine, and isoleucine), and charged (arginine) side chains and methylated pyrimidinic (1-methylcytosine and 1-methylthymine) and puric (9-methyladenine and 9-methylguanine) DNA bases. The computational approach combined different methodologies: a molecular mechanics (MMFFs forced field) conformational search and structural and vibrational density-functional calculations (M06-2X with double and triple-ζ Pople's basis sets). To dissect the interactions, intermolecular forces were analyzed with the Non-Covalent Interactions (NCI) analysis. The results for the 24 different clusters studied show a noticeable correlation between the calculated binding energies and the propensities for protein-DNA base interactions found in the literature. Such correlation holded even for the interaction of the selected amino acid derivatives with Watson and Crick pairs. Therefore, the balance between hydrogen bonds and van der Waals interactions (specially stacking) in the control of the final shape of the investigated amino acid-DNA base pairs seems to be well reproduced in dispersion-corrected DFT molecular models, reinforcing the idea that the specificity between the amino acids and the DNA bases play an important role in the regulation of DNA. PMID:26765058

  2. Diverse levels of sequence selectivity and catalytic efficiency of protein-tyrosine phosphatases.

    PubMed

    Selner, Nicholas G; Luechapanichkul, Rinrada; Chen, Xianwen; Neel, Benjamin G; Zhang, Zhong-Yin; Knapp, Stefan; Bell, Charles E; Pei, Dehua

    2014-01-21

    The sequence selectivity of 14 classical protein-tyrosine phosphatases (PTPs) (PTPRA, PTPRB, PTPRC, PTPRD, PTPRO, PTP1B, SHP-1, SHP-2, HePTP, PTP-PEST, TCPTP, PTPH1, PTPD1, and PTPD2) was systematically profiled by screening their catalytic domains against combinatorial peptide libraries. All of the PTPs exhibit similar preference for pY peptides rich in acidic amino acids and disfavor positively charged sequences but differ vastly in their degrees of preference/disfavor. Some PTPs (PTP-PEST, SHP-1, and SHP-2) are highly selective for acidic over basic (or neutral) peptides (by >10(5)-fold), whereas others (PTPRA and PTPRD) show no to little sequence selectivity. PTPs also have diverse intrinsic catalytic efficiencies (kcat/KM values against optimal substrates), which differ by >10(5)-fold due to different kcat and/or KM values. Moreover, PTPs show little positional preference for the acidic residues relative to the pY residue. Mutation of Arg47 of PTP1B, which is located near the pY-1 and pY-2 residues of a bound substrate, decreased the enzymatic activity by 3-18-fold toward all pY substrates containing acidic residues anywhere within the pY-6 to pY+5 region. Similarly, mutation of Arg24, which is situated near the C-terminus of a bound substrate, adversely affected the kinetic activity of all acidic substrates. A cocrystal structure of PTP1B bound with a nephrin pY(1193) peptide suggests that Arg24 engages in electrostatic interactions with acidic residues at the pY+1, pY+2, and likely other positions. These results suggest that long-range electrostatic interactions between positively charged residues near the PTP active site and acidic residues on pY substrates allow a PTP to bind acidic substrates with similar affinities, and the varying levels of preference for acidic sequences by different PTPs are likely caused by the different electrostatic potentials near their active sites. The implications of the varying sequence selectivity and intrinsic catalytic

  3. Fatty acid profiles in relation to triglyceride level in the liver of dairy cows.

    PubMed

    Sato, Hiroshi; Mohamed, Tharwat; Goto, Akiko; Oikawa, Shin; Kurosawa, Takashi

    2004-01-01

    To elucidate possible relationships between triglyceride (TG) levels and fatty acid composition in bovine liver, hepatic TG and seven individual fatty acids were measured in 23 Holstein dairy cows, of them 6 are healthy. Liver TG level was greater than 3 % in 12 cows which were ruled fatty liver. Palmitic and oleic acid proportions were significantly higher in fatty liver cows than in the healthy cows, while stearic acid was lower in fatty liver cows. With increased liver TG, stearic acid proportions decreased dramatically. Results indicate that hepatic lipidosis markedly alters the proportions of the various fatty acids in the liver of dairy cows.

  4. An experimental and theoretical study of the amino acid side chain Raman bands in proteins

    NASA Astrophysics Data System (ADS)

    Sjöberg, Béatrice; Foley, Sarah; Cardey, Bruno; Enescu, Mironel

    2014-07-01

    The Raman spectra of a series of tripeptides with the basic formula GlyAAGly where the central amino acid (AA) was tryptophan, tyrosine, phenylalanine, glycine, methionine, histidine, lysine and leucine were measured in H2O. The theoretical Raman spectra obtained using density functional theory (DFT) calculations at the B3LYP/6-311+G(2df,2pd) level of theory allows a precise attribution of the vibrational bands. The experimental results show that there is a blue shift in the frequencies of several bands of the amino acid side chains in tripeptides compared to free amino acids, especially in the case of AAs containing aromatic rings. On the other hand, a very good agreement was found between the Raman bands of AA residues in tripeptides and those measured on three model proteins: bovine serum albumin, β-lactoglobulin and lysozyme. The present analysis contributes to an unambiguous interpretation of the protein Raman spectra that is useful in monitoring the biological reactions involving AA side chains alteration.

  5. Proteins of calcified endoskeleton: II partial amino acid sequences of endoskeletal proteins and the characterization of proteinaceous organic matrix of spicules from the alcyonarian, Synularia polydactyla.

    PubMed

    Rahman, M Azizur; Isa, Yeishin; Uehara, Tsuyoshi

    2005-03-01

    Calcified organic substances in the skeleton contain a protein-polysaccharide complex taking a key role in the regulation of bio-calcification. However, information concerning the matrix proteins in alcyonarian and their effect on calcification process is still unknown. For this reason, we have studied the organic matrix of endoskeletal spicules from the alcyonarian coral, Synularia polydactyla, to analyze the proteins with their sequences and investigate the functional properties by a molecular approach. The separated spicules from the colony were identified by scanning electron microscope (SEM). The soluble organic matrix comprised 0.04% of spicule weight. By recording decline of pH in the experimental design, the inhibitory effect of the matrix on CaCO3 precipitation was revealed. Prior to electrophoresis, our analysis of proteins extracted from the soluble organic matrix of the spicules revealed an abundance of proteins in molecular weight. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the preparations showed seven bands of proteins with an apparent molecular mass of 109, 83, 70, 63, 41, 30 and 22 kDa. The proteins were electrophoresed on Tricine-SDS-PAGE after electro-elution treatment, and then transferred to polyvinylidene difluoride (PVDF) membranes and their N-termini were sequenced. Two bands of proteins of about 70 and 63 kDa successfully underwent N-terminal amino acid sequencing. For the detection of calcium binding proteins, a Ca2+ overlay analysis was conducted on the extract by 45Ca autoradiography. The 109 and 63 kDa calcium binding proteins were found to be radioactive. Periodic acid schiff staining indicated that 83 and 63 kDa proteins were glycosylated. An assay for carbonic anhydrase, which is thought to play an important role in the process of calcification revealed low level of the activity. These findings suggest that the endoskeletal spicules of alcyonarian corals have protein-rich organic matrices

  6. Monocyte chemoattractant protein-1 serum levels in ovarian cancer patients

    PubMed Central

    Hefler, L; Tempfer, C; Heinze, G; Mayerhofer, K; Breitenecker, G; Leodolter, S; Reinthaller, A; Kainz, C

    1999-01-01

    The chemokine monocyte chemoattractant protein (MCP)-1 is an important mediator of monocyte infiltration in various solid tumours of epithelial origin. The aim of the present study was to evaluate the role of MCP-1 in the natural history of ovarian cancer and to determine its value as differentiation marker and prognostic marker regarding disease free and overall survival. This retrospective study comprises 86 patients with ovarian cancer, 48 with primary ovarian cancer and 38 with recurrent ovarian cancer, 67 patients with benign ovarian cysts and 42 healthy women. Median serum levels in patients with primary ovarian cancer, recurrent ovarian cancer, benign ovarian cysts and in healthy women were 535.6 (range 129.6–1200) pg ml–1, 427.3 (range 193.4–1101) pg ml–1, 371.2 (range 222–986.8) pg ml–1 and 318.7 (range 241.3–681.4) pg ml–1 respectively (Mann–Whitney U-test, P < 0.001). Univariate logistic regression models revealed a significant influence of MCP-1 serum levels on the odds of presenting with primary ovarian cancer versus benign cysts and versus healthy women respectively (univariate logistic regression, P < 0.001 and P < 0.001 respectively). In a multivariate logistic regression model considering MCP-1 and CA 125 serum levels simultaneously, both MCP-1 and CA 125 revealed statistical significance on the odds of presenting with primary ovarian cancer versus benign cysts (multivariate logistic regression, P = 0.05 and P < 0.001 respectively). In ovarian cancer patients, MCP-1 serum levels showed a statistically significant correlation with histological grade (Mann–Whitney U-test, P = 0.02) and age at the time of diagnosis (Mann–Whitney U-test, P = 0.03). Elevated MCP-1 serum levels prior to therapy were not associated with disease-free and overall survival (log-rank test, P = 0.2 and P = 0.7 respectively). In summary these data indicate that MCP-1 might play a functional role in the natural history of ovarian cancer and might serve as

  7. Plasma Free Fatty Acids, Fatty Acid-binding Protein 4, and Mortality in Older Adults (From the Cardiovascular Health Study)

    PubMed Central

    Miedema, Michael D.; Maziarz, Marlena; Biggs, Mary L.; Zieman, Susan J.; Kizer, Jorge R.; Ix, Joachim H.; Mozaffarian, Dariush; Tracy, Russell P.; Psaty, Bruce M.; Siscovick, David S.; Mukamal, Kenneth J.; Djousse, Luc

    2014-01-01

    Plasma free fatty acids (FFA) are largely derived from adipose tissue. Elevated levels of FFA and fatty acid-binding protein 4 (FABP4), a key cytoplasmic chaperone of fatty acids, have been associated with adverse cardiovascular outcomes but limited data are available on the relation of these biomarkers with cardiovascular and total mortality. We studied 4,707 participants with a mean age of 75 years who had plasma FFA and FABP4 measured in 1992–1993 as part of the Cardiovascular Health Study, an observational cohort of community dwelling older adults. Over a median follow-up of 11.8 years, 3,555 participants died. Cox proportional hazard regression was used to determine the association between FFA, FABP4, and mortality. In fully adjusted models, FFA were associated with dose-dependent significantly higher total mortality (hazard ratio (HR) per standard deviation (SD): 1.14, 95% confidence interval (CI) 1.09–1.18), but FABP4 levels were not (HR 1.04, 95% CI 0.98–1.09). In a cause-specific mortality analysis, higher concentrations of FFA were associated with significantly higher risk of death due to cardiovascular disease, dementia, infection, and respiratory causes, but not cancer or trauma. We did not find evidence of an interaction between FFA and FABP4 (p=0.45), but FABP4 appeared to be associated with total mortality differentially among men and women (HR 1.17 (1.08–1.26) for men, HR 1.02 (0.96–1.07) for women, interaction p-value <0.001). In conclusion, in a cohort of community-dwelling older individuals, elevated plasma concentrations of FFA, but not FABP4, were associated with cardiovascular and non-cardiovascular mortality. PMID:25073566

  8. Site specific incorporation of heavy atom-containing unnatural amino acids into proteins for structure determination

    DOEpatents

    Xie, Jianming; Wang, Lei; Wu, Ning; Schultz, Peter G.

    2008-07-15

    Translation systems and other compositions including orthogonal aminoacyl tRNA-synthetases that preferentially charge an orthogonal tRNA with an iodinated or brominated amino acid are provided. Nucleic acids encoding such synthetases are also described, as are methods and kits for producing proteins including heavy atom-containing amino acids, e.g., brominated or iodinated amino acids. Methods of determining the structure of a protein, e.g., a protein into which a heavy atom has been site-specifically incorporated through use of an orthogonal tRNA/aminoacyl tRNA-synthetase pair, are also described.

  9. Influence of dietary protein type and iron source on the absorption of amino acids and minerals.

    PubMed

    Pérez-Llamas, F; Garaulet, M; Martínez, J A; Marín, J F; Larqué, E; Zamora, S

    2001-12-01

    The apparent digestibility coefficient (ADC) of amino acids and the balance of minerals (calcium, phosphorus, magnesium and iron) has been determined in rats fed four diets differing in the protein type (casein or soy protein) and iron source (ferrous sulphate or lactate) in order to study the possible interactions of these nutrients. The availability of amino acids, especially essential amino acids, was greater in the diet made with animal protein (casein). The iron source also affected the absorption of most amino acids in all the diets assayed with ferrous sulphate being greater. The balance of iron, magnesium and phosphorus was higher in the diets containing animal protein. The retention of calcium and magnesium was significantly greater when ferrous sulphate was used as iron source. These results demonstrate the important interaction between amino acids and minerals and between the minerals themselves, which must be carefully studied when selecting different types of protein or mineral sources in human or animal nutrition.

  10. Systematic Identification of Single Amino Acid Variants in Glioma Stem-Cell-Derived Chromosome 19 Proteins

    PubMed Central

    2015-01-01

    Novel proteoforms with single amino acid variations represent proteins that often have altered biological functions but are less explored in the human proteome. We have developed an approach, searching high quality shotgun proteomic data against an extended protein database, to identify expressed mutant proteoforms in glioma stem cell (GSC) lines. The systematic search of MS/MS spectra using PEAKS 7.0 as the search engine has recognized 17 chromosome 19 proteins in GSCs with altered amino acid sequences. The results were further verified by manual spectral examination, validating 19 proteoforms. One of the novel findings, a mutant form of branched-chain aminotransferase 2 (p.Thr186Arg), was verified at the transcript level and by targeted proteomics in several glioma stem cell lines. The structure of this proteoform was examined by molecular modeling in order to estimate conformational changes due to mutation that might lead to functional modifications potentially linked to glioma. Based on our initial findings, we believe that our approach presented could contribute to construct a more complete map of the human functional proteome. PMID:25399873

  11. Interaction of perfluoroalkyl acids with human liver fatty acid-binding protein.

    PubMed

    Sheng, Nan; Li, Juan; Liu, Hui; Zhang, Aiqian; Dai, Jiayin

    2016-01-01

    Perfluoroalkyl acids (PFAAs) are highly persistent and bioaccumulative, resulting in their broad distribution in humans and the environment. The liver is an important target for PFAAs, but the mechanisms behind PFAAs interaction with hepatocyte proteins remain poorly understood. We characterized the binding of PFAAs to human liver fatty acid-binding protein (hL-FABP) and identified critical structural features in their interaction. The binding interaction of PFAAs with hL-FABP was determined by fluorescence displacement and isothermal titration calorimetry (ITC) assay. Molecular simulation was conducted to define interactions at the binding sites. ITC measurement revealed that PFOA/PFNA displayed a moderate affinity for hL-FABP at a 1:1 molar ratio, a weak binding affinity for PFHxS and no binding for PFHxA. Moreover, the interaction was mainly mediated by electrostatic attraction and hydrogen bonding. Substitution of Asn111 with Asp caused loss of binding affinity to PFAA, indicating its crucial role for the initial PFAA binding to the outer binding site. Substitution of Arg122 with Gly caused only one molecule of PFAA to bind to hL-FABP. Molecular simulation showed that substitution of Arg122 increased the volume of the outer binding pocket, making it impossible to form intensive hydrophobic stacking and hydrogen bonds with PFOA, and highlighting its crucial role in the binding process. The binding affinity of PFAAs increased significantly with their carbon number. Arg122 and Asn111 played a pivotal role in these interactions. Our findings may help understand the distribution pattern, bioaccumulation, elimination, and toxicity of PFAAs in humans.

  12. A case for protein-level and site-level specificity in glycoproteomic studies of disease.

    PubMed

    Schumacher, Katherine N; Dodds, Eric D

    2016-06-01

    Abnormal glycosylation of proteins is known to be either resultant or causative of a variety of diseases. This makes glycoproteins appealing targets as potential biomarkers and focal points of molecular studies on the development and progression of human ailment. To date, a majority of efforts in disease glycoproteomics have tended to center on either determining the concentration of a given glycoprotein, or on profiling the total population of glycans released from a mixture of glycoproteins. While these approaches have demonstrated some diagnostic potential, they are inherently insensitive to the fine molecular detail which distinguishes unique and possibly disease relevant glycoforms of specific proteins. As a consequence, such analyses can be of limited sensitivity, specificity, and accuracy because they do not comprehensively consider the glycosylation status of any particular glycoprotein, or of any particular glycosylation site. Therefore, significant opportunities exist to improve glycoproteomic inquiry into disease by engaging in these studies at the level of individual glycoproteins and their exact loci of glycosylation. In this concise review, the rationale for glycoprotein and glycosylation site specificity is developed in the context of human disease glycoproteomics with an emphasis on N-glycosylation. Recent examples highlighting disease-related perturbations in glycosylation will be presented, including those involving alterations in the overall glycosylation of a specific protein, alterations in the occupancy of a given glycosylation site, and alterations in the compositional heterogeneity of glycans occurring at a given glycosylation site. Each will be discussed with particular emphasis on how protein-specific and site-specific approaches can contribute to improved discrimination between glycoproteomes and glycoproteins associated with healthy and unhealthy states.

  13. Uses of phage display in agriculture: sequence analysis and comparative modeling of late embryogenesis abundant client proteins suggest protein-nucleic acid binding functionality.

    PubMed

    Kushwaha, Rekha; Downie, A Bruce; Payne, Christina M

    2013-01-01

    A group of intrinsically disordered, hydrophilic proteins-Late Embryogenesis Abundant (LEA) proteins-has been linked to survival in plants and animals in periods of stress, putatively through safeguarding enzymatic function and prevention of aggregation in times of dehydration/heat. Yet despite decades of effort, the molecular-level mechanisms defining this protective function remain unknown. A recent effort to understand LEA functionality began with the unique application of phage display, wherein phage display and biopanning over recombinant Seed Maturation Protein homologs from Arabidopsis thaliana and Glycine max were used to retrieve client proteins at two different temperatures, with one intended to represent heat stress. From this previous study, we identified 21 client proteins for which clones were recovered, sometimes repeatedly. Here, we use sequence analysis and homology modeling of the client proteins to ascertain common sequence and structural properties that may contribute to binding affinity with the protective LEA protein. Our methods uncover what appears to be a predilection for protein-nucleic acid interactions among LEA client proteins, which is suggestive of subcellular residence. The results from this initial computational study will guide future efforts to uncover the protein protective mechanisms during heat stress, potentially leading to phage-display-directed evolution of synthetic LEA molecules.

  14. Influence of Bleaching on Flavor of 34% Whey Protein Concentrate and Residual Benzoic Acid Concentration in Dried Whey Proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies have shown that bleaching negatively affects the flavor of 70% whey protein concentrate (WPC70), but bleaching effects on lower-protein products have not been established. Benzoyl peroxide (BP), a whey bleaching agent, degrades to benzoic acid (BA) and may elevate BA concentrations...

  15. Dormancy alleviation by NO or HCN leading to decline of protein carbonylation levels in apple (Malus domestica Borkh.) embryos.

    PubMed

    Krasuska, Urszula; Ciacka, Katarzyna; Dębska, Karolina; Bogatek, Renata; Gniazdowska, Agnieszka

    2014-08-15

    Deep dormancy of apple (Malus domestica Borkh.) embryos can be overcome by short-term pre-treatment with nitric oxide (NO) or hydrogen cyanide (HCN). Dormancy alleviation of embryos modulated by NO or HCN and the first step of germination depend on temporary increased production of reactive oxygen species (ROS). Direct oxidative attack on some amino acid residues or secondary reactions via reactive carbohydrates and lipids can lead to the formation of protein carbonyl derivatives. Protein carbonylation is a widely accepted covalent and irreversible modification resulting in inhibition or alteration of enzyme/protein activities. It also increases the susceptibility of proteins to proteolytic degradation. The aim of this work was to investigate protein carbonylation in germinating apple embryos, the dormancy of which was removed by pre-treatment with NO or HCN donors. It was performed using a quantitative spectrophotometric method, while patterns of carbonylated protein in embryo axes were analyzed by immunochemical techniques. The highest concentration of protein carbonyl groups was observed in dormant embryos. It declined in germinating embryos pre-treated with NO or HCN, suggesting elevated degradation of modified proteins during seedling formation. A decrease in the concentration of carbonylated proteins was accompanied by modification in proteolytic activity in germinating apple embryos. A strict correlation between the level of protein carbonyl groups and cotyledon growth and greening was detected. Moreover, direct in vitro carbonylation of BSA treated with NO or HCN donors was analyzed, showing action of both signaling molecules as protein oxidation agents. PMID:24973585

  16. Dormancy alleviation by NO or HCN leading to decline of protein carbonylation levels in apple (Malus domestica Borkh.) embryos.

    PubMed

    Krasuska, Urszula; Ciacka, Katarzyna; Dębska, Karolina; Bogatek, Renata; Gniazdowska, Agnieszka

    2014-08-15

    Deep dormancy of apple (Malus domestica Borkh.) embryos can be overcome by short-term pre-treatment with nitric oxide (NO) or hydrogen cyanide (HCN). Dormancy alleviation of embryos modulated by NO or HCN and the first step of germination depend on temporary increased production of reactive oxygen species (ROS). Direct oxidative attack on some amino acid residues or secondary reactions via reactive carbohydrates and lipids can lead to the formation of protein carbonyl derivatives. Protein carbonylation is a widely accepted covalent and irreversible modification resulting in inhibition or alteration of enzyme/protein activities. It also increases the susceptibility of proteins to proteolytic degradation. The aim of this work was to investigate protein carbonylation in germinating apple embryos, the dormancy of which was removed by pre-treatment with NO or HCN donors. It was performed using a quantitative spectrophotometric method, while patterns of carbonylated protein in embryo axes were analyzed by immunochemical techniques. The highest concentration of protein carbonyl groups was observed in dormant embryos. It declined in germinating embryos pre-treated with NO or HCN, suggesting elevated degradation of modified proteins during seedling formation. A decrease in the concentration of carbonylated proteins was accompanied by modification in proteolytic activity in germinating apple embryos. A strict correlation between the level of protein carbonyl groups and cotyledon growth and greening was detected. Moreover, direct in vitro carbonylation of BSA treated with NO or HCN donors was analyzed, showing action of both signaling molecules as protein oxidation agents.

  17. Human skeletal muscle and erythrocyte proteins involved in acid-base homeostasis: adaptations to chronic hypoxia.

    PubMed

    Juel, C; Lundby, C; Sander, M; Calbet, J A L; Hall, G van

    2003-04-15

    Chronic hypoxia is accompanied by changes in blood and skeletal muscle acid-base control. We hypothesized that the underlying mechanisms include altered protein expression of transport systems and the enzymes involved in lactate, HCO3- and H+ fluxes in skeletal muscle and erythrocytes. Immunoblotting was used to quantify densities of the transport systems and enzymes. Muscle and erythrocyte samples were obtained from eight Danish lowlanders at sea level and after 2 and 8 weeks at 4100 m (Bolivia). For comparison, samples were obtained from eight Bolivian natives. In muscle membranes there were no changes in fibre-type distribution, lactate dehydrogenase isoforms, Na+,K+-pump subunits or in the lactate-H+ co-transporters MCT1 and MCT4. The Na+-H+ exchanger protein NHE1 was elevated by 39 % in natives compared to lowlanders. The Na+-HCO3- co-transporter density in muscle was elevated by 47-69 % after 2 and 8 weeks at altitude. The membrane-bound carbonic anhydrase (CA) IV in muscle increased in the lowlanders by 39 %, whereas CA XIV decreased by 23-47 %. Levels of cytosolic CA II and III in muscle and CA I and II in erythrocytes were unchanged. The erythrocyte lactate-H+ co-transporter MCT1 increased by 230-405 % in lowlanders and was 324 % higher in natives. The erythrocyte inorganic anion exchanger (Cl--HCO3- exchanger AE1) was increased by 149-228 %. In conclusion, chronic hypoxia induces dramatic changes in erythrocyte proteins, but only moderate changes in muscle proteins involved in acid-base control. Together, these changes suggest a hypoxia-induced increase in the capacity for lactate, HCO3- and H+ fluxes from muscle to blood and from blood to erythrocytes. PMID:12611920

  18. Hepatic fatty acid biosynthesis is more responsive to protein than carbohydrate in rainbow trout during acute stimulations.

    PubMed

    Dai, Weiwei; Panserat, Stéphane; Kaushik, Sadasivam; Terrier, Frédéric; Plagnes-Juan, Elisabeth; Seiliez, Iban; Skiba-Cassy, Sandrine

    2016-01-01

    The link between dietary carbohydrate/protein and de novo lipogenesis (DNL) remains debatable in carnivorous fish. We aimed to evaluate and compare the response of hepatic lipogenic gene expression to dietary carbohydrate intake/glucose and dietary protein intake/amino acids (AAs) during acute stimulations using both in vivo and in vitro approaches. For the in vivo trial, three different diets and a controlled-feeding method were employed to supply fixed amount of dietary protein or carbohydrate in a single meal; for the in vitro trial, primary hepatocytes were stimulated with a low or high level of glucose (3 mM or 20 mM) and a low or high level of AAs (one-fold or four-fold concentrated AAs). In vitro data showed that a high level of AAs upregulated the expression of enzymes involved in DNL [fatty acid synthase (FAS) and ATP citrate lyase (ACLY)], lipid bioconversion [elongation of very long chain fatty acids like-5 (Elovl5), Elovl2, Δ6 fatty acyl desaturase (D6D) and stearoyl-CoA desaturase-1 (SCD1)], NADPH production [glucose-6-phosphate dehydrogenase (G6PDH) and malic enzyme (ME)], and transcriptional factor sterol regulatory element binding protein 1-like, while a high level of glucose only elevated the expression of ME. Data in trout liver also showed that high dietary protein intake induced higher lipogenic gene expression (FAS, ACLY, and Elovl2) regardless of dietary carbohydrate intake, while high carbohydrate intake markedly suppressed the expression of acetyl-CoA carboxylase (ACC) and Elovl5. Overall, we conclude that, unlike rodents or humans, hepatic fatty acid biosynthetic gene expression in rainbow trout is more responsive to dietary protein intake/AAs than dietary carbohydrate intake/glucose during acute stimulations. This discrepancy probably represents one important physiological and metabolic difference between carnivores and omnivores. PMID:26491101

  19. Hepatic fatty acid biosynthesis is more responsive to protein than carbohydrate in rainbow trout during acute stimulations.

    PubMed

    Dai, Weiwei; Panserat, Stéphane; Kaushik, Sadasivam; Terrier, Frédéric; Plagnes-Juan, Elisabeth; Seiliez, Iban; Skiba-Cassy, Sandrine

    2016-01-01

    The link between dietary carbohydrate/protein and de novo lipogenesis (DNL) remains debatable in carnivorous fish. We aimed to evaluate and compare the response of hepatic lipogenic gene expression to dietary carbohydrate intake/glucose and dietary protein intake/amino acids (AAs) during acute stimulations using both in vivo and in vitro approaches. For the in vivo trial, three different diets and a controlled-feeding method were employed to supply fixed amount of dietary protein or carbohydrate in a single meal; for the in vitro trial, primary hepatocytes were stimulated with a low or high level of glucose (3 mM or 20 mM) and a low or high level of AAs (one-fold or four-fold concentrated AAs). In vitro data showed that a high level of AAs upregulated the expression of enzymes involved in DNL [fatty acid synthase (FAS) and ATP citrate lyase (ACLY)], lipid bioconversion [elongation of very long chain fatty acids like-5 (Elovl5), Elovl2, Δ6 fatty acyl desaturase (D6D) and stearoyl-CoA desaturase-1 (SCD1)], NADPH production [glucose-6-phosphate dehydrogenase (G6PDH) and malic enzyme (ME)], and transcriptional factor sterol regulatory element binding protein 1-like, while a high level of glucose only elevated the expression of ME. Data in trout liver also showed that high dietary protein intake induced higher lipogenic gene expression (FAS, ACLY, and Elovl2) regardless of dietary carbohydrate intake, while high carbohydrate intake markedly suppressed the expression of acetyl-CoA carboxylase (ACC) and Elovl5. Overall, we conclude that, unlike rodents or humans, hepatic fatty acid biosynthetic gene expression in rainbow trout is more responsive to dietary protein intake/AAs than dietary carbohydrate intake/glucose during acute stimulations. This discrepancy probably represents one important physiological and metabolic difference between carnivores and omnivores.

  20. Milk production responses to dietary stearic acid vary by production level in dairy cattle.

    PubMed

    Piantoni, P; Lock, A L; Allen, M S

    2015-03-01

    Effects of stearic acid supplementation on feed intake and metabolic and production responses of dairy cows with a wide range of milk production (32.2 to 64.4 kg/d) were evaluated in a crossover design experiment with a covariate period. Thirty-two multiparous Holstein cows (142±55 d in milk) were assigned randomly within level of milk yield to treatment sequence. Treatments were diets supplemented (2% of diet dry matter) with stearic acid (SA; 98% C18:0) or control (soyhulls). The diets were based on corn silage and alfalfa and contained 24.5% forage neutral detergent fiber, 25.1% starch, and 17.3% crude protein. Treatment periods were 21 d with the final 4 d used for data and sample collection. Compared with the control, SA increased dry matter intake (DMI; 26.1 vs. 25.2 kg/d) and milk yield (40.2 vs. 38.5 kg/d). Stearic acid had no effect on the concentration of milk components but increased yields of fat (1.42 vs. 1.35 kg/d), protein (1.19 vs. 1.14 kg/d), and lactose (1.96 vs. 1.87 kg/d). The SA treatment increased 3.5% fat-corrected milk (3.5% FCM; 40.5 vs. 38.6 kg/d) but did not affect feed efficiency (3.5% FCM/DMI, 1.55 vs. 1.53), body weight, or body condition score compared with the control. Linear interactions between treatment and level of milk yield during the covariate period were detected for DMI and yields of milk, fat, protein, lactose, and 3.5% FCM; responses to SA were positively related to milk yield of cows. The SA treatment increased crude protein digestibility (67.4 vs. 65.5%), tended to increase neutral detergent fiber digestibility (43.6 vs. 42.3%), decreased fatty acid (FA) digestibility (56.6 vs. 76.1%), and did not affect organic matter digestibility. Fatty acid yield response, calculated as the additional FA yield secreted in milk per unit of additional FA intake, was only 13.3% for total FA and 8.2% for C18:0 plus cis-9 C18:1. Low estimated digestibility of the SA supplement was at least partly responsible for the low FA yield response

  1. Allied Health Chemistry Laboratory: Amino Acids, Insulin, Proteins, and Skin

    ERIC Educational Resources Information Center

    Dever, David F.

    1975-01-01

    Presents a laboratory experiment specifically designed for allied health students. The students construct molecular models of amino acids, extract amino acids from their skin with hot water, and chromatographically analyze the skin extract and hydrolyzed insulin. (MLH)

  2. The human ubiquitin-52 amino acid fusion protein gene shares several structural features with mammalian ribosomal protein genes.

    PubMed Central

    Baker, R T; Board, P G

    1991-01-01

    Complementary DNA clones encoding ubiquitin fused to a 52 amino acid tail protein were isolated from human placental and adrenal gland cDNA libraries. The deduced human 52 amino acid tail protein is very similar to the homologous protein from other species, including the conservation of the putative metal-binding, nucleic acid-binding domain observed in these proteins. Northern blot analysis with a tail-specific probe indicated that the previously identified UbA mRNA species most likely represents comigrating transcripts of the 52 amino acid tail (UbA52) and 80 amino acid tail (UbA80) ubiquitin fusion genes. The UbA52 gene was isolated from a human genomic library and consists of five exons distributed over 3400 base pairs. One intron is in the 5' non-coding region, two interrupt the single ubiquitin coding unit, and the fourth intron is within the tail coding region. Several members of the Alu family of repetitive DNA are associated with the gene. The UbA52 promoter has several features in common with mammalian ribosomal protein genes, including its location in a CpG-rich island, initiation of transcription within a polypyrimidine tract, the lack of a consensus TATA motif, and the presence of Sp1 binding sites, observations that are consistent with the recent identification of the ubiquitin-free tail proteins as ribosomal proteins. Thus, in spite of its unusual feature of being translationally fused to ubiquitin, the 52 amino acid tail ribosomal protein is expressed from a structurally typical ribosomal protein gene. Images PMID:1850507

  3. Assessment of Folic Acid Supplementation in Pregnant Women by Estimation of Serum Levels of Tetrahydrofolic Acid, Dihydrofolate Reductase, and Homocysteine.

    PubMed

    Naithani, Manisha; Saxena, Vartika; Mirza, Anissa Atif; Kumari, Ranjeeta; Sharma, Kapil; Bharadwaj, Jyoti

    2016-01-01

    Background. Status of folic acid use in pregnant women of the hilly regions in North India was little known. This study was carried out to assess the folic acid use and estimate folate metabolites in pregnant women of this region. Materials and Methods. This cross-sectional study is comprised of 76 pregnant women, whose folic acid supplementation was assessed by a questionnaire and serum levels of homocysteine, tetrahydrofolic acid (THFA), and dihydrofolate reductase (DHFR) were estimated using Enzyme Linked Immunoassays. Results. The study data revealed awareness of folic acid use during pregnancy was present in 46.1% and 23.7% were taking folic acid supplements. The study depicted that there was no statistically significant difference between serum levels of THFA and DHFR in pregnant women with and without folic acid supplements (p = 0.790). Hyperhomocysteinemia was present in 15.78% of the participants. Conclusion. Less awareness about folic acid supplementation and low use of folic acid by pregnant women were observed in this region. Sufficient dietary ingestion may suffice for the escalated requirements in pregnancy, but since this cannot be ensured, hence folic acid supplementation should be made as an integral part of education and reproductive health programs for its better metabolic use, growth, and development of fetus.

  4. Assessment of Folic Acid Supplementation in Pregnant Women by Estimation of Serum Levels of Tetrahydrofolic Acid, Dihydrofolate Reductase, and Homocysteine

    PubMed Central

    Saxena, Vartika; Mirza, Anissa Atif; Kumari, Ranjeeta; Sharma, Kapil; Bharadwaj, Jyoti

    2016-01-01

    Background. Status of folic acid use in pregnant women of the hilly regions in North India was little known. This study was carried out to assess the folic acid use and estimate folate metabolites in pregnant women of this region. Materials and Methods. This cross-sectional study is comprised of 76 pregnant women, whose folic acid supplementation was assessed by a questionnaire and serum levels of homocysteine, tetrahydrofolic acid (THFA), and dihydrofolate reductase (DHFR) were estimated using Enzyme Linked Immunoassays. Results. The study data revealed awareness of folic acid use during pregnancy was present in 46.1% and 23.7% were taking folic acid supplements. The study depicted that there was no statistically significant difference between serum levels of THFA and DHFR in pregnant women with and without folic acid supplements (p = 0.790). Hyperhomocysteinemia was present in 15.78% of the participants. Conclusion. Less awareness about folic acid supplementation and low use of folic acid by pregnant women were observed in this region. Sufficient dietary ingestion may suffice for the escalated requirements in pregnancy, but since this cannot be ensured, hence folic acid supplementation should be made as an integral part of education and reproductive health programs for its better metabolic use, growth, and development of fetus. PMID:27064332

  5. Abscisic acid and late embryogenesis abundant protein profile changes in winter wheat under progressive drought stress.

    PubMed

    Vaseva, I I; Grigorova, B S; Simova-Stoilova, L P; Demirevska, K N; Feller, U

    2010-09-01

    Three varieties (cv. Pobeda, Katya and Sadovo) of winter wheat (Triticum aestivum), differing in their agronomic characteristics, were analysed during progressive soil water stress and recovery at early vegetation stages. Changes in abscisic acid content, SDS-PAGE and immunoblot profiles of proteins that remained soluble upon heating were monitored. Initially higher ABA content in control Pobeda and Katya corresponded to earlier expression of the studied late embryogenesis abundant (LEA) proteins. A combination of higher ABA content, early immunodetection of dehydrins, and a significant increase of WZY2 transcript levels were observed in drought-stressed leaves of the tolerant variety Katya. One-step RT-PCR analyses of some acidic dehydrin genes (WCOR410b, TADHN) documented their relatively constant high expression levels in leaves under drought stress during early vegetative development. Neutral WZY2 dehydrin, TaLEA2 and TaLEA3 transcripts accumulated gradually with increasing water deficit. Delayed expression of TaLEA2 and TaLEA3 genes was found in the least drought-tolerant wheat, Sadovo. The expression profile of WZY2 revealed two distinct and separate bands, suggesting alternative splicing, which altered as water stress increased.

  6. Plastidial fatty acid levels regulate resistance gene-dependent defense signaling in Arabidopsis.

    PubMed

    Chandra-Shekara, A C; Venugopal, Srivathsa C; Barman, Subhankar Roy; Kachroo, Aardra; Kachroo, Pradeep

    2007-04-24

    In Arabidopsis, resistance to Turnip Crinkle Virus (TCV) depends on the resistance (R) gene, HRT, and the recessive locus rrt. Resistance also depends on salicylic acid (SA), EDS1, and PAD4. Exogenous application of SA confers resistance in RRT-containing plants by increasing HRT transcript levels in a PAD4-dependent manner. Here we report that reduction of oleic acid (18:1) can also induce HRT gene expression and confer resistance to TCV. However, the 18:1-regulated pathway is independent of SA, rrt, EDS1, and PAD4. Reducing the levels of 18:1, via a mutation in the SSI2-encoded stearoyl-acyl carrier protein-desaturase, or by exogenous application of glycerol, increased transcript levels of HRT as well as several other R genes. Second-site mutations in the ACT1-encoded glycerol-3-phosphate acyltransferase or GLY1-encoded glycerol-3-phosphate dehydrogenase restored 18:1 levels in HRT ssi2 plants and reestablished a dependence on rrt. Resistance to TCV and HRT gene expression in HRT act1 plants was inducible by SA but not by glycerol, whereas that in HRT pad4 plants was inducible by glycerol but not by SA. The low 18:1-mediated induction of R gene expression was also dependent on ACT1 but independent of EDS1, PAD4, and RAR1. Intriguingly, TCV inoculation did not activate this 18:1-regulated pathway in HRT plants, but instead resulted in the induction of several genes that encode 18:1-synthesizing isozymes. These results suggest that the 18:1-regulated pathway may be specifically targeted during pathogen infection and that altering 18:1 levels may serve as a unique strategy for promoting disease resistance.

  7. Ubiquitin/proteasome pathway regulates levels of retinoic acid receptor gamma and retinoid X receptor alpha in human keratinocytes.

    PubMed

    Boudjelal, M; Wang, Z; Voorhees, J J; Fisher, G J

    2000-04-15

    Repeated exposure of human skin to solar UV radiation leads to premature aging (photoaging) and skin cancer. UV-induced skin damage can be ameliorated by all-trans retinoic acid treatment. The actions of retinoic acid in skin keratinocytes are mediated primarily by nuclear retinoic acid receptor gamma (RARgamma) and retinoid X receptor alpha (RXRalpha). We found that exposure of cultured primary human keratinocytes to UV irradiation (30 mJ/cm2) substantially reduced (50-90%) RARgamma and RXRalpha mRNA and protein within 8 h. The rates of disappearance of RARgamma and RXRalpha proteins after UV exposure or treatment with the protein synthesis inhibitor cycloheximide were similar. UV irradiation did not increase the rate of breakdown of RARgamma or RXRalpha but rather reduced their rate of synthesis. The addition of proteasome inhibitors MG132 and LLvL, but not the lysosomal inhibitor E64, prevented loss of RARgamma and RXRalpha proteins after exposure of keratinocytes to either UV radiation or cycloheximide. Soluble extracts from nonirradiated or UV-irradiated keratinocytes possessed similar levels of proteasome activity that degraded RARgamma and RXRalpha proteins in vitro. Furthermore, RARgamma and RXRalpha were polyubiquitinated in intact cells. RXRalpha was found to contain two proline, glutamate/aspartate, serine, and threonine (PEST) motifs, which confer rapid turnover of many short-lived regulatory proteins that are degraded by the ubiquitin/proteasome pathway. However, the PEST motifs in RXRalpha did not function to regulate its stability, because deletion of the PEST motifs individually or together did not alter ubiquitination or proteasome-mediated degradation of RXRalpha. These results demonstrate that loss of RARgamma and RXRalpha proteins after UV irradiation results from degradation via the ubiquitin/proteasome pathway. Taken together, the data here indicate that ubiquitin/proteasome-mediated breakdown is an important mechanism regulating the levels of

  8. Amino acid rating method for evaluating protein adequacy of infant formulas.

    PubMed

    Sarwar, G; Botting, H G; Peace, R W

    1989-01-01

    Amino acid profiles and/or protein digestibility (by the rat balance method) were determined for various forms (powder, ready-to-use, liquid concentrate, etc.) of cow's milk- and soy-based infant formulas obtained from 4 manufacturers. The essential amino acid data of the formulas were compared with that of human milk for the calculation of amino acid scores (based on the single most limiting amino acid). The product of amino acid score and total protein (g/100 kcal) was then termed "amino acid rating." Amino acid scores for the milk- and soy-based formulas ranged from 59 to 90 and from 59 to 81%, respectively, due to deficiencies in sulfur amino acids and/or tryptophan. Because of significantly higher total protein contents (g/100 kcal) of soy- (2.65-3.68) and milk-based (2.20-2.95) infant formulas compared to human milk (1.5), the relative amino acid ratings (human milk = 100) for all infant formulas except 2 liquid concentrates (having values of 87%) were above 100%. Values for true digestibility of protein in milk- and soy-based formulas ranged from 87 to 97 and from 92 to 95%, respectively. When corrected for protein digestibility, the relative amino acid ratings for all the milk-based liquid concentrates were below 100% (77-98%).

  9. Biophysical and computational methods to analyze amino acid interaction networks in proteins.

    PubMed

    O'Rourke, Kathleen F; Gorman, Scott D; Boehr, David D

    2016-01-01

    Globular proteins are held together by interacting networks of amino acid residues. A number of different structural and computational methods have been developed to interrogate these amino acid networks. In this review, we describe some of these methods, including analyses of X-ray crystallographic data and structures, computer simulations, NMR data, and covariation among protein sequences, and indicate the critical insights that such methods provide into protein function. This information can be leveraged towards the design of new allosteric drugs, and the engineering of new protein function and protein regulation strategies. PMID:27441044

  10. Genetically programmed expression of proteins containing the unnatural amino acid phenylselenocysteine

    DOEpatents

    Wang, Jiangyun; Schultz, Peter G.

    2010-09-07

    The invention relates to orthogonal pairs of tRNAs and aminoacyl-tRNA synthetases that can incorporate the unnatural amino acid phenylselenocysteine into proteins produced in eubacterial host cells such as E. coli. The invention provides, for example but not limited to, novel orthogonal aminoacyl-tRNA synthetases, polynucleotides encoding the novel synthetase molecules, methods for identifying and making the novel synthetases, methods for producing proteins containing the unnatural amino acid phenylselenocysteine and translation systems. The invention further provides methods for producing modified proteins (e.g., lipidated proteins) through targeted modification of the phenylselenocysteine residue in a protein.

  11. Genetically programmed expression of proteins containing the unnatural amino acid phenylselenocysteine

    DOEpatents

    Wang, Jiangyun; Schultz, Peter G.

    2012-07-10

    The invention relates to orthogonal pairs of tRNAs and aminoacyl-tRNA synthetases that can incorporate the unnatural amino acid phenylselenocysteine into proteins produced in eubacterial host cells such as E. coli. The invention provides, for example but not limited to, novel orthogonal aminoacyl-tRNA synthetases, polynucleotides encoding the novel synthetase molecules, methods for identifying and making the novel synthetases, methods for producing proteins containing the unnatural amino acid phenylselenocysteine and translation systems. The invention further provides methods for producing modified proteins (e.g., lipidated proteins) through targeted modification of the phenylselenocysteine residue in a protein.

  12. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut (Juglans regia L.) proteins and protein fractionations.

    PubMed

    Mao, Xiaoying; Hua, Yufei; Chen, Guogang

    2014-01-27

    As a by-product of oil production, walnut proteins are considered as an additional source of plant protein for human food. To make full use of the protein resource, a comprehensive understanding of composition and characteristics of walnut proteins are required. Walnut proteins have been fractionated and characterized in this study. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut proteins and protein fractionations were analyzed. The proteins were sequentially separated into four fractions according to their solubility. Glutelin was the main component of the protein extract. The content of glutelin, albumin, globulin and prolamin was about 72.06%, 7.54%, 15.67% and 4.73% respectively. Glutelin, albumin and globulin have a balanced content of essential amino acids, except for methionine, with respect to the FAO pattern recommended for adults. SDS-PAGE patterns of albumin, globulin and glutelin showed several polypeptides with molecular weights 14.4 to 66.2 kDa. The pattern of walnut proteins in two-dimension electrophoresis (2-DE) showed that the isoelectric point was mainly in the range of 4.8-6.8. The results of size exclusion chromatogram indicated molecular weight of the major components of walnut proteins were between 3.54 and 81.76 kDa.

  13. Amino Acid Composition, Molecular Weight Distribution and Gel Electrophoresis of Walnut (Juglans regia L.) Proteins and Protein Fractionations

    PubMed Central

    Mao, Xiaoying; Hua, Yufei; Chen, Guogang

    2014-01-01

    As a by-product of oil production, walnut proteins are considered as an additional source of plant protein for human food. To make full use of the protein resource, a comprehensive understanding of composition and characteristics of walnut proteins are required. Walnut proteins have been fractionated and characterized in this study. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut proteins and protein fractionations were analyzed. The proteins were sequentially separated into four fractions according to their solubility. Glutelin was the main component of the protein extract. The content of glutelin, albumin, globulin and prolamin was about 72.06%, 7.54%, 15.67% and 4.73% respectively. Glutelin, albumin and globulin have a balanced content of essential amino acids, except for methionine, with respect to the FAO pattern recommended for adults. SDS-PAGE patterns of albumin, globulin and glutelin showed several polypeptides with molecular weights 14.4 to 66.2 kDa. The pattern of walnut proteins in two-dimension electrophoresis (2-DE) showed that the isoelectric point was mainly in the range of 4.8–6.8. The results of size exclusion chromatogram indicated molecular weight of the major components of walnut proteins were between 3.54 and 81.76 kDa. PMID:24473146

  14. Identification of thermophilic proteins by incorporating evolutionary and acid dissociation information into Chou's general pseudo amino acid composition.

    PubMed

    Fan, Guo-Liang; Liu, Yan-Ling; Wang, Hui

    2016-10-21

    Thermophilic proteins can thrive stalely at the high temperatures. Identification of thermophilic protein could be helpful to learn the function of protein. Automated prediction of thermophilic protein is an important tool for genome annotation. In this work, a powerful predictor is proposed by combining amino acid composition, evolutionary information, and acid dissociation constant. The overall prediction accuracy of 93.53% was obtained for using the algorithm of support vector machine. In order to check the performance of our method, two low-similarity independent testing datasets are used to test the proposed method. Comparisons with other methods show that the prediction results were better than other existing methods in literature. This indicates that our approach was effective to predict thermophilic proteins. PMID:27396359

  15. Oral supplementation with whey proteins increases plasma glutathione levels of HIV-infected patients.

    PubMed

    Micke, P; Beeh, K M; Schlaak, J F; Buhl, R

    2001-02-01

    HIV infection is characterized by an enhanced oxidant burden and a systemic deficiency of the tripeptide glutathione (GSH), a major antioxidant. The semi-essential amino acid cysteine is the main source of the free sulfhydryl group of GSH and limits its synthesis. Therefore, different strategies to supplement cysteine supply have been suggested to increase glutathione levels in HIV-infected individuals. The aim of this study was to evaluate the effect of oral supplementation with two different cysteine-rich whey protein formulas on plasma GSH levels and parameters of oxidative stress and immune status in HIV-infected patients. In a prospective double blind clinical trial, 30 patients (25 male, 5 female; mean age (+/- SD) 42 +/- 9.8 years) with stable HIV infection (221 +/- 102 CD4 + lymphocytes L-1) were randomized to a supplemental diet with a daily dose of 45 g whey proteins of either Protectamin (Fresenius Kabi, Bad Hamburg, Germany) or Immunocal (Immunotec, Vandreuil, Canada) for two weeks. Plasma concentrations of total, reduced and oxidized GSH, superoxide anion (O2-) release by blood mononuclear cells, plasma levels of TNF-alpha and interleukins 2 and 12 were quantified with standard methods at baseline and after therapy. Pre-therapy, plasma GSH levels (Protectamin: 1.92 +/- 0.6 microM; Immunocal: 1.98 +/- 0.9 microM) were less than normal (2.64 +/- 0.7 microM, P = 0.03). Following two weeks of oral supplementation with whey proteins, plasma GSH levels increased in the Protectamin group by 44 +/- 56% (2.79 +/- 1.2 microM, P = 0.004) while the difference in the Immunocal group did not reach significance (+ 24.5 +/- 59%, 2.51 +/- 1.48 microM, P = 0.43). Spontaneous O2- release by blood mononuclear cells was stable (20.1 +/- 14.2 vs. 22.6 +/- 16.1 nmol h-1 10-6 cells, P = 0.52) whereas PMA-induced O2- release decreased in the Protectamin group (53.7 +/- 19 vs. 39.8 +/- 18 nmol h-1 10-6 cells, P = 0.04). Plasma concentrations of TNF-alpha and interleukins 2 and

  16. Amino acid sequence of the encephalitogenic basic protein from human myelin

    PubMed Central

    Carnegie, P. R.

    1971-01-01

    Myelin from the central nervous system contains an unusual basic protein, which can induce experimental autoimmune encephalomyelitis. The basic protein from human brain was digested with trypsin and other enzymes and the sequence of the 170 amino acids was determined. The localization of the encephalitogenic determinants was described. Possible roles for the protein in the structure and function of myelin are discussed. PMID:4108501

  17. Hypertonic stress regulates amino acid transport and cell cycle proteins in chick embryo hepatocytes.

    PubMed

    Bruscalupi, Giovannella; Massimi, Mara; Spagnuolo, Silvana; Fiore, Anna Maria; Leoni, Silvia

    2012-02-01

    Hyperosmotic stress affects cell growth, decreasing cell volume and increasing the uptake of organic osmolytes. However, the sensitivity of embryonic cells to osmotic treatment remains to be established. We have analysed some aspects of cell-cycle control and amino-acid transport in hypertonic conditions during prenatal life. The effects of hyperosmotic stress on amino-acid uptake mediated by system A, (3)H-thymidine incorporation, and regulation of cell-cycle proteins were analysed in chick embryo hepatocytes. Hypertonic stress increased system A activity and caused cell-cycle delay. Effects on amino-acid transport involved p38 kinase activation and new carrier synthesis. Cyclin D1, cdk4 (cyclin-dependent kinase 4) and PCNA (proliferating-cell nuclear antigen) levels decreased, whereas cyclin E, p21 and p53 levels were unchanged. Incorporation of (3)H-leucine indicated decreased synthesis of cyclin D1. In contrast, analysis of mRNA by qRT-PCR (quantitative real-time PCR) showed a net increase of cyclin D1 transcripts, suggesting post-transcriptional regulation. The data show that chick embryo hepatocytes respond to hyperosmotic conditions by arresting cell growth to prevent DNA damage and increasing osmolyte uptake to regulate cell volume, indicating that the adaptive response to environmental stress exists during prenatal life.

  18. TMV-Cg Coat Protein stabilizes DELLA proteins and in turn negatively modulates salicylic acid-mediated defense pathway during Arabidopsis thaliana viral infection

    PubMed Central

    2014-01-01

    Background Plant viral infections disturb defense regulatory networks during tissue invasion. Emerging evidence demonstrates that a significant proportion of these alterations are mediated by hormone imbalances. Although the DELLA proteins have been reported to be central players in hormone cross-talk, their role in the modulation of hormone signaling during virus infections remains unknown. Results This work revealed that TMV-Cg coat protein (CgCP) suppresses the salicylic acid (SA) signaling pathway without altering defense hormone SA or jasmonic acid (JA) levels in Arabidopsis thaliana. Furthermore, it was observed that the expression of CgCP reduces plant growth and delays the timing of floral transition. Quantitative RT-qPCR analysis of DELLA target genes showed that CgCP alters relative expression of several target genes, indicating that the DELLA proteins mediate transcriptional changes produced by CgCP expression. Analyses by fluorescence confocal microscopy showed that CgCP stabilizes DELLA proteins accumulation in the presence of gibberellic acid (GA) and that the DELLA proteins are also stabilized during TMV-Cg virus infections. Moreover, DELLA proteins negatively modulated defense transcript profiles during TMV-Cg infection. As a result, TMV-Cg accumulation was significantly reduced in the quadruple-DELLA mutant Arabidopsis plants compared to wild type plants. Conclusions Taken together, these results demonstrate that CgCP negatively regulates the salicylic acid-mediated defense pathway by stabilizing the DELLA proteins during Arabidopsis thaliana viral infection, suggesting that CgCP alters the stability of DELLAs as a mechanism of negative modulation of antiviral defense responses. PMID:25084837

  19. Modulation of sialic acid levels among some organs during insulin resistance or hyperglycemic states.

    PubMed

    Ibrahim, Mohammed Auwal; Abdulkadir, Aisha; Onojah, Alice; Sani, Lawal; Adamu, Auwal; Abdullahi, Hadiza

    2016-01-01

    Previous studies have suggested a possible connection between insulin resistance and chronic hyperglycemia with membrane sialic acid content. In this study, the effects of high (20% ad libitum) fructose and glucose feeding on the sialic acid levels of some organs were investigated in rats. The blood glucose levels of the high fructose- and glucose-fed rats were consistently and significantly (P < 0.05) higher than the normal control throughout the experiment. Free serum sialic acid and total hepatic sialic acid levels were elevated in the high fructose- and glucose-fed rats compared to normal control, but only the data for the high glucose-fed group were significantly (P < 0.05) different from the normal control. Conversely, a significant (P < 0.05) decrease in the pancreatic sialic acid level was observed in high glucose-fed group compared to normal control. Also, the high fructose-fed rats had lower, but insignificant (P > 0.05), pancreatic sialic acid level than the normal control. On the other hand, high fructose and glucose feeding did not significantly (P > 0.05) affect the sialic acid levels of the skeletal muscle and heart, though a tendency to increase the sialic acid level was evident in the heart. In the kidney, the sialic acid level was significantly (P < 0.05) increased in both high fructose- and glucose-fed groups. It was concluded that the liver and kidney tend to stimulate sialic acid synthesis, while the pancreas downregulate sialic acids synthesis and/or promote sialic acid release from glycoconjugates. Also, these organs may contribute to high-serum sialic acid level observed during diabetes. PMID:26468092

  20. Lipoic acid entrains the hepatic circadian clock and lipid metabolic proteins that have been desynchronized with advanced age

    SciTech Connect

    Keith, Dove; Finlay, Liam; Butler, Judy; Gómez, Luis; Smith, Eric; Moreau, Régis; Hagen, Tory

    2014-07-18

    Highlights: • 24 month old rats were supplemented with 0.2% lipoic acid in the diet for 2 weeks. • Lipoic acid shifts phase of core circadian clock proteins. • Lipoic acid corrects age-induced desynchronized lipid metabolism rhythms. - Abstract: It is well established that lipid metabolism is controlled, in part, by circadian clocks. However, circadian clocks lose temporal precision with age and correlates with elevated incidence in dyslipidemia and metabolic syndrome in older adults. Because our lab has shown that lipoic acid (LA) improves lipid homeostasis in aged animals, we hypothesized that LA affects the circadian clock to achieve these results. We fed 24 month old male F344 rats a diet supplemented with 0.2% (w/w) LA for 2 weeks prior to sacrifice and quantified hepatic circadian clock protein levels and clock-controlled lipid metabolic enzymes. LA treatment caused a significant phase-shift in the expression patterns of the circadian clock proteins Period (Per) 2, Brain and Muscle Arnt-Like1 (BMAL1), and Reverse Erythroblastosis virus (Rev-erb) β without altering the amplitude of protein levels during the light phase of the day. LA also significantly altered the oscillatory patterns of clock-controlled proteins associated with lipid metabolism. The level of peroxisome proliferator-activated receptor (PPAR) α was significantly increased and acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) were both significantly reduced, suggesting that the LA-supplemented aged animals are in a catabolic state. We conclude that LA remediates some of the dyslipidemic processes associated with advanced age, and this mechanism may be at least partially through entrainment of circadian clocks.

  1. The Influence of Tobacco Smoke on Protein and Metal Levels in the Serum of Women during Pregnancy

    PubMed Central

    Wrześniak, Marta; Kepinska, Marta; Królik, Małgorzata; Milnerowicz, Halina

    2016-01-01

    Background Tobacco smoking by pregnant women has a negative effect on fetal development and increases pregnancy risk by changing the oxidative balance and microelements level. Smoking affects the concentration, structure and function of proteins, potentially leading to various negative effects on pregnancy outcomes. Methodology/Principal Findings The influence of tobacco smoke on key protein fractions in smoking and non-smoking healthy pregnant women was determined by capillary electrophoresis (CE). Concentrations of the proteins α1-antitrypsin, α1-acid glycoprotein, α2-macroglobulin and transferrin were determined by ELISA tests. Total protein concentration was measured by the Biuret method. Smoking status was established by cotinine levels. Cadmium (Cd) and Zinc (Zn) concentrations were determined by flame atomic absorption spectrometry and the Zn/Cd ratio was calculated based on these numbers. Smoking women had a 3.7 times higher level of Cd than non-smoking women. Zn levels decreased during pregnancy for all women. The Zn/Cd ratio was three times lower in smoking women. The differences between the changes in the protein profile for smoking and non-smoking women were noted. Regarding proteins, α1-antitrypsin and α2-macroglobulin levels were lower in the non-smoking group than in the smoking group and correlated with Cd levels (r = -0.968, p = 0.032 for non-smokers; r = −0.835, p = 0.019 for smokers). Zn/Cd ratios correlated negatively with α1-, α2- and β-globulins. Conclusions/Significance Exposure to tobacco smoke increases the concentration of Cd in the blood of pregnant women and may lead to an elevated risk of pregnancy disorders. During pregnancy alter concentrations of some proteins. The correlation of Cd with proteins suggests that it is one of the causes of protein aberrations. PMID:27548057

  2. [L-type fatty acid binding protein (L-FABP) and kidney disease].

    PubMed

    Kamijo-Ikemori, Atsuko; Sugaya, Takeshi; Kimura, Kenjiro

    2014-02-01

    Liver-type fatty acid binding protein (L-FABP) is expressed in the cytoplasm of human renal proximal tubules. Renal L-FABP expression is up-regulated and urinary excretion of renal L-FABP is increased by various stressors, such as urinary protein, hyperglycemia, tubular ischemia, toxins, and salt-sensitive hypertension, which lead to the progression of kidney disease. Urinary L-FABP levels accurately reflect the degree of tubulointerstitial damage and are strongly correlated with the prognosis of chronic kidney disease (CKD) patients in clinical studies. In patients with type I or type II diabetes, urinary L-FABP levels were reported to be significantly higher in patients with normal levels of urinary albumin than in those with microalbuminuria. Urinary L-FABP may be useful for the early detection of diabetic nephropathy. Furthermore, in a longitudinal study, a higher level of urinary L-FABP was found to be a risk factor for the progression of diabetic nephropathy. With respect to acute kidney disease (AKI), urinary L-FABP facilitates the early detection of AKI before an increase in serum creatinine. Therefore, urinary L-FABP was approved as a new tubular biomarker by the Ministry of Health, Labour and Welfare of Japan.

  3. Amino acid metabolism, substrate availability and the control of protein dynamics in the human kidney.

    PubMed

    Garibotto, G; Tessari, P; Sacco, P; Deferrari, G

    1999-01-01

    The mechanisms controlling protein metabolism in the human kidney are not well understood. During adult life, kidney protein content and the size of the kidney remain fairly constant, indicating that protein synthesis and degradation within the kidney are tightly regulated. However, kidney protein turnover may change in response to stimuli such as alterations in substrate availability, hormones or growth factors, acid-base balance, renal work or renal injury with a progressive decrease in the number of nephrons. These factors have been evaluated mainly in animals, in vitro or in vivo. Amino acids, the kidneys substrates for protein synthesis, are provided by several routes. Like in other organs, amino acids can reach the kidney cells through the arterial blood flow. However, they may also come from the degradation of reabsorbed low-molecular weight proteins filtered by the glomerulus. The human kidney has high rates of protein turnover and leucine oxidation. The magnitude of the protein turnover across the human kidney suggests that the protein dynamics is partly determined by intrarenal protein catabolism. As evaluated by a steady-state leucine multiple compartment analysis, kidney protein synthesis is dependent to a similar extent on intrarenal generation of amino acids from protein breakdown and from amino acids taken up from the arterial blood. Kidney mass may therefore depend not only on the availability of free amino acids, but also on filtered proteins which are degraded within the kidney. Future studies could define the mechanisms, metabolic pathways and mediators influencing kidney protein turnover in humans, with a view to better comprehension of the mechanisms of disease. PMID:10493563

  4. [Antinociceptive effect of docosahexaenoic acid (DHA) through long fatty acid receptor G protein-coupled receptor 40 (GPR40)].

    PubMed

    Nakamoto, Kazuo; Nishinaka, Takashi; Sato, Naoya; Mankura, Mitsumasa; Koyama, Yutaka; Tokuyama, Shogo

    2014-01-01

    Fatty acids, one class of essential nutrients for humans, are an important source of energy and an essential component of cell membranes. They also function as signal transduction molecules in a variety of biological phenomena. The important functional role of fatty acids in both onset and suppression of pain has become increasingly apparent in recent years. Recently, we have also demonstrated that the release of an endogenous opioid peptide, β-endorphin, plays an important role in the induction of docosahexaenoic acid (DHA)-induced antinociception. It is well known that fatty acids affect intracellular and intercellular signaling as well as the membrane fluidity of neurons. In addition to intracellular actions, unbound free fatty acids (FFAs) can also carry out extracellular signaling by stimulating the G protein-coupled receptor (GPCR). Among these receptors, G protein-coupled receptor 40 (GPR40) has been reported to be activated by long-chain fatty acids such as DHA, eicosapentaenoic acid (EPA) and arachidonic acid. In the peripheral area, GPR40 is preferentially expressed in pancreatic β-cells and is known to relate to the secretion of hormone and peptides. On the other hand, even though this receptor is widely distributed in the central nervous system, reports studying the role and functions of GPR40 in the brain have not been found. In this review, we summarize the findings of our recent study about the long-chain fatty acid receptor GPR40 as a novel pain regulatory system. PMID:24584021

  5. Fast computational methods for predicting protein structure from primary amino acid sequence

    DOEpatents

    Agarwal, Pratul Kumar

    2011-07-19

    The present invention provides a method utilizing primary amino acid sequence of a protein, energy minimization, molecular dynamics and protein vibrational modes to predict three-dimensional structure of a protein. The present invention also determines possible intermediates in the protein folding pathway. The present invention has important applications to the design of novel drugs as well as protein engineering. The present invention predicts the three-dimensional structure of a protein independent of size of the protein, overcoming a significant limitation in the prior art.

  6. High levels of stearic acid, palmitoleic acid, and dihomo-γ-linolenic acid and low levels of linoleic acid in serum cholesterol ester are associated with high insulin resistance.

    PubMed

    Kurotani, Kayo; Sato, Masao; Ejima, Yuko; Nanri, Akiko; Yi, Siyan; Pham, Ngoc Minh; Akter, Shamima; Poudel-Tandukar, Kalpana; Kimura, Yasumi; Imaizumi, Katsumi; Mizoue, Tetsuya

    2012-09-01

    The association of fatty acid composition with insulin resistance and type 2 diabetes has been reported in Western populations, but there is limited evidence of this association among the Japanese, whose populace consume large amounts of fish. To test the hypothesis that high palmitic, palmitoleic, and dihomo-γ-linolenic acids and low levels of linoleic and n-3 fatty acids are associated with higher insulin resistance among the Japanese, the authors investigated the relationship between serum fatty acid composition and serum C-peptide concentrations in 437 Japanese employees aged 21 to 67 years who participated in a workplace health examination. Serum cholesterol ester and phospholipid fatty acid compositions were measured by gas-liquid chromatography. Desaturase activity was estimated by fatty acid product-to-precursor ratios. A multiple regression was used to assess the association between fatty acid and C-peptide concentrations. C-peptide concentrations were associated inversely with linoleic acid levels in cholesterol ester and phospholipid (P for trend = .01 and .02, respectively) and positively with stearic and palmitoleic acids in cholesterol ester (P for trend =.02 and .006, respectively) and dihomo-γ-linolenic acid in cholesterol ester and phospholipid (P for trend < .0001 for both). C-peptide concentrations were not associated with n-3 polyunsaturated fatty acids. C-peptide concentrations significantly increased as δ-9-desaturase (16:1 n-7/16:0) and δ-6-desaturase (18:3 n-6/18:2 n-6) increased (P for trend = .01 and .03, respectively) and δ-5-desaturase (20:4 n-6/20:3 n-6) decreased (P for trend = .004). In conclusion, a fatty acid pattern with high levels of serum stearic, palmitoleic, or dihomo-γ-linolenic acids; δ-9-desaturase (16:1 n-7/16:0) or δ-6-desaturase (18:3 n-6/18:2 n-6) activities; and low levels of serum linoleic acid or δ-5-desaturase (20:4 n-6/20:3 n-6) activity might be associated with higher insulin resistance in Japanese adults.

  7. Detection of D-amino acids in purified proteins synthesized in Escherichia coli.

    PubMed

    Miyamoto, Tetsuya; Sekine, Masae; Ogawa, Tetsuhiro; Hidaka, Makoto; Homma, Hiroshi; Masaki, Haruhiko

    2010-05-01

    It has long been believed that amino acids comprising proteins of all living organisms are only of the L-configuration, except for Gly. However, peptidyl D-amino acids were observed in hydrolysates of soluble high molecular weight fractions extracted from cells or tissues of various organisms. This strongly suggests that significant amounts of D-amino acids are naturally present in usual proteins. Thus we analyzed the D-amino acid contents of His-tag-purified beta-galactosidase and human urocortin, which were synthesized by Escherichia coli grown in controlled synthetic media. After acidic hydrolysis for various times at 110 degrees C, samples were derivatized with 4-fluoro-7-nitro-2, 1, 3-benzoxadiazole (NBD-F) and separated on a reverse-phase column followed by a chiral column into D- and L-enantiomers. The contents of D-enantiomers of Ala, Leu, Phe, Val, Asp, and Glu were determined by plotting index D/(D + L) against the incubation time for hydrolysis and extrapolating the linear regression line to 0 h to eliminate the effect of racemization of amino acids during the incubation. Significant contents of D-amino acids were reproducibly detected, the D-amino acid profile being specific to an individual protein. This finding indicated the likelihood that D-amino acids are in fact present in the purified proteins. On the other hand, the D-amino acid contents of proteins were hardly influenced by the addition of D- or L-amino acids to the cultivation medium, whereas intracellular free D-amino acids sensitively varied according to the extracellular conditions. The origin of these D-amino acids detected in proteins was discussed.

  8. Evaluation of postmortem urea nitrogen, creatinine and uric acid levels in pericardial fluid in forensic autopsy.

    PubMed

    Zhu, Bao-Li; Ishikawa, Takaki; Michiue, Tomomi; Li, Dong-Ri; Zhao, Dong; Quan, Li; Maeda, Hitoshi

    2005-10-01

    In postmortem biochemistry, there is insufficient data available for the practical analysis of factors in the pericardial fluid. The aim of the present study was to examine postmortem pericardial fluid for urea nitrogen (UN), creatinine (Cr) and uric acid (UA) levels to investigate the pathophysiology of death in forensic autopsy cases (total, n = 409; within 48 h postmortem), which included blunt, sharp instrument injury, asphyxiation, drowning, fire fatalities, hyperthermia, hypothermia, methamphetamine-related fatalities, other poisoning, delayed death from trauma and natural diseases. There was a significant elevation in the three markers for chronic renal failure, gastrointestinal bleeding, hyperthermia, hypothermia, methamphetamine fatalities and delayed traumatic death, which was comparable with the clinical criteria for their serum levels. These postmortem findings showed azotemia due to renal failure, elevated protein catabolism and rhabdomyolysis. Although the pericardial levels were otherwise similar to the clinical serum reference ranges, only the drowning fatalities showed significantly lower levels for each marker. These observations suggested the stability of UN, Cr and UA in the pericardial fluid within 48 h postmortem and their usefulness for the pathophysiological investigation of death involving azotemia.

  9. Sulforaphane reduces the alterations induced by quinolinic acid: modulation of glutathione levels.

    PubMed

    Santana-Martínez, R A; Galván-Arzáte, S; Hernández-Pando, R; Chánez-Cárdenas, M E; Avila-Chávez, E; López-Acosta, G; Pedraza-Chaverrí, J; Santamaría, A; Maldonado, P D

    2014-07-11

    Glutamate-induced excitotoxicity involves a state of acute oxidative stress, which is a crucial event during neuronal degeneration and is part of the physiopathology of neurodegenerative diseases. In this work, we evaluated the ability of sulforaphane (SULF), a natural dietary isothiocyanate, to induce the activation of transcription factor Nrf2 (a master regulator of redox state in the cell) in a model of striatal degeneration in rats infused with quinolinic acid (QUIN). Male Wistar rats received SULF (5mg/kg, i.p.) 24h and 5min before the intrastriatal infusion of QUIN. SULF increased the reduced glutathione (GSH) levels 4h after QUIN infusion, which was associated with its ability to increase the activity of glutathione reductase (GR), an antioxidant enzyme capable to regenerate GSH levels at 24h. Moreover, SULF treatment increased glutathione peroxidase (GPx) activity, while no changes were observed in γ-glutamyl cysteine ligase (GCL) activity. SULF treatment also prevented QUIN-induced oxi